51-52-5KNAHARQHSZJURB-UHFFFAOYSA-NKNAHARQHSZJURB-UHFFFAOYSA-N
6-Propyl-2-thiouracil6-Propyl-2 thiouracil (PTU)
4(1H)-Pyrimidinone, 2,3-dihydro-6-propyl-2-thioxo-
2,3-Dihydro-6-propyl-2-thioxo-4(1H)-pyrimidinone
2-Mercapto-4-hydroxy-6-n-propylpyrimidine
2-Mercapto-4-hydroxy-6-propylpyrimidine
2-Mercapto-6-propylpyrimidin-4-ol
2-Thio-4-oxo-6-propyl-1,3-pyrimidine
2-Thio-6-propyl-1,3-pyrimidin-4-one
6-n-Propyl-2-thiouracil
6-n-Propylthiouracil
6-Propyl-2-thio-2,4(1H,3H)pyrimidinedione
6-Propylthiouracil
NSC 6498
NSC 70461
Procasil
Propacil
propiltiouracilo
Propycil
Propyl-Thiorist
Propylthiorit
propylthiouracil
Propylthiouracile
Propyl-Thyracil
Prothiucil
Prothiurone
Prothycil
Prothyran
Protiural
Thiuragyl
Thyreostat II
URACIL, 4-PROPYL-2-THIO-
Uracil, 6-propyl-2-thio-
DTXSID502120960-56-0PMRYVIKBURPHAH-UHFFFAOYSA-NPMRYVIKBURPHAH-UHFFFAOYSA-N
Methimazole2H-Imidazole-2-thione, 1,3-dihydro-1-methyl-
1,3-Dihydro-1-methyl-2H-imidazole-2-thione
1-Methyl-1,3-dihydroimidazole-2-thione
1-Methyl-1H-imidazole-2-thiol
1-Methyl-2-mercapto-1H-imidazole
1-Methyl-2-mercaptoimidazole
1-Methyl-4-imidazoline-2-thione
1-Methylimidazole-2(3H)-thione
1-Methylimidazole-2-thiol
1-Methylimidazole-2-thione
2-Mercapto-1-methyl-1H-imidazole
2-Mercapto-1-methylimidazole
2-Mercapto-N-methylimidazole
4-Imidazoline-2-thione, 1-methyl-
Basolan
Danantizol
Favistan
Frentirox
Imidazole-2-thiol, 1-methyl-
Mercaptazole
Mercazole
Mercazolyl
Metazolo
Methimazol
Methylmercaptoimidazole
Metothyrin
Metothyrine
Metotirin
N-Methyl-2-mercaptoimidazole
N-Methylimidazolethiol
NSC 38608
Strumazol
Tapazole
Thacapzol
Thiamazol
thiamazole
Thycapzol
Thymidazol
Thymidazole
tiamazol
DTXSID4020820UBERON:0002421hippocampal formationGO:0001508action potentialGO:0007420brain developmentGO:0007612learningGO:0007613memory9disrupted8morphological change2decreasedPyrethrins and Pyrethroids2016-11-29T18:42:272016-11-29T18:42:27Propylthiouracil2016-11-29T18:42:222016-11-29T18:42:22Methimazole2016-11-29T18:42:192016-11-29T18:42:19WikiUser_28VertebratesWikiUser_29Invertebrates10090mouse10116ratWCS_9606humanWCS_7227fruit flyWCS_7955zebrafishWCS_160004gastropodsBinding to voltage-gated sodium channelBinding to VGSCMolecular<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Ion channels are integral membrane proteins that are critical for neuronal function. They form pores in the plasma membrane that allow certain ions to travel with their concentration gradient across the membrane. Those that open in response to a change in membrane voltage potential are called voltage-gated ion channels. Channels that open in response to binding using a chemical signal or molecule are ligand-gated ion channels. In neurons, ion channels are essential for chemical communication between cells, or synaptic transmission. Ion channels also function to maintain membrane potential and initiate and propagate electrical impulses. Voltage-gated sodium channels are therefore responsible for action potential initiation and propagation in excitable cells, including nerve, muscle and neuroendocrine cell types. They are also expressed at low levels in non-excitable cells. It is important to note is that functional VGSC are present in both grey and white matter in the brain and approximately 50% of white matter oligodendrocyte precursor cells producing trains of action potentials and receiving synaptic input (Fields, 2008). VGSC are also present on microglia cells and contribute to release of major pro-inflammatory cytokines (Hossain et al., 2017).</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Mammalian VGSC are composed of one α and two β subunits. Ten separate α subunits (Ogata and Ohishi, 2002) and four different β subunits (Isom, 2002) have been identified and are expressed in a tissue, region and time specific manner. The diverse functional roles of VGSCs depend on the numerous potential combinations of α and β subunits (Ogata and Ohishi, 2002). The type of VGSCs expressed in different cell types and regions, and their sensitivity and their functional role, may all contribute to the manifestation of toxicity and age dependent sensitivity, including the effects caused by pyrethroids.</span></span></p>
<p style="text-align:justify"><img alt="" src="https://aopwiki.org/system/dragonfly/production/2022/03/15/9bkwiiz0mg_Picture1.jpg" /></p>
<p><span style="font-size:8pt"><span style="font-family:Tahoma,sans-serif">At resting membrane potentials the channel is closed. During the rising phase of an action potential the channel activates or opens. Channel inactivation contributes to the falling phase. During the undershoot phase the channel deactivates before returning to the closed phase once resting membrane resting potential has been restored. Source: adapted from Motifolio Biomedical PowerPoint Toolkit Suite.</span></span></p>
<p style="text-align:justify"> </p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">The sodium channel protein has been discovered and characterised in biochemical and molecular detail, even to atomic resolution. The initial works performed to measure and detect the electrical signals in nerves were initiated by Hodgkin and Huxley in 1952, showing a voltage‐dependent activation of sodium current that carries Na<sup>+</sup> inward. The structure of VGSCs is nowadays known in detail and some seminal papers are available (Catterall, 2012).</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Intracellular microelectrode recording using voltage or patch clamp are the common methods used for electrophysiological studies of VGSC. Channels and locations can also be measured using immunohistochemical methods, transcriptomics and at protein levels.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Expression of different sodium channel isoforms can be measured using a panel of sodium channel subunit-specific antibodies. Quantification of immunocytochemical staining is difficult due to differences in equipment, tissue preparation, inter-assay variability and analysis methods. However, using a quantitative approach, it is possible to determine the localisation and relative levels of sodium channel subunit protein expression (Westenbroek et al., 2013). PCR amplification and competitive PCR approach, real-time PCR, are used to isolate the mRNA levels of VGSC isoforms (Haufe et al., 2005).</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Every cell within living organisms actively maintains an intracellular Na<sup>+</sup> concentration that is 10–12 times lower than the extracellular concentration. The cells then utilise this transmembrane Na<sup>+</sup> concentration gradient as a driving force to produce electrical signals, sometimes in the form of action potentials. The protein family comprising VGSC (Na<sub>v</sub>s) is essential for such signalling and enables cells to change their status in a regenerative manner and to rapidly communicate with one another. VGSC were first predicted in squid and were later identified through molecular biology in the electric eel. Since then, these proteins have been discovered in organisms ranging from bacteria to humans (Chaihne, 2018). </span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Sodium channels consist of a highly processed α subunit, which is approximately 260 kDa, associated with auxiliary β subunits of 33–39 kDa. Sodium channels in the adult central nervous system (CNS) and heart contain a mixture of β1–β4 subunits, while sodium channels in adult skeletal muscle have only the β1 subunit. Nine different sodium channels have been identified using electrophysiological recording, biochemical purification, and cloning (Catterall, 2012).</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Nomenclature of the different sodium channels utilises a numerical system to define subfamilies and subtypes based on similarities between the amino acid sequences of the channels. In this nomenclature system, the name of an individual channel consists of the chemical symbol of the principal permeating ion (Na) with the principal physiological regulator (voltage) indicated as a subscript (Na<sub>v</sub>). The number following the subscript indicates the gene subfamily (currently only Na<sub>v </sub>1), and the number following the full point identifies the specific channel isoform (e.g. Na­<sub>v</sub> 1.1). This last number has been assigned according to the approximate order in which each gene was identified. Splice variants of each family member are identified by lower-case letters following the numbers (e.g. Na<sub>v</sub> 1.1a). Nine mammalian sodium channel isoforms have been identified and functionally expressed with all greater than 50% identical in amino acid sequence in the transmembrane and extracellular domains, where the amino acid sequence is similar enough for clear alignment (Catterall, 2012). In addition to these nine sodium channels that have been functionally expressed, closely related sodium channel-like proteins (Nax) have been cloned from mouse, rat and human. They are approximately 50% identical to the Na<sub>v</sub> 1 subfamily of channels but more than 80% identical to each other (Catterall, 2012).</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">In mammals, neuronal VGSC are expressed in the adult and developing brain. Evidence from mutation and knockout animal models demonstrates that perturbation of VGSC function during development impair nervous system structure and function, including muscle function, pain reception and cardia arrythmias (Chahine, 2018). VGSC show complex regional and temporal ontogeny in mammals (see <strong>Table 1</strong>, from Shafer et al., 2005). In general, embryonically expressed forms of VGSCs are replaced by expression of adult forms as neurodevelopment proceeds.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><img alt="" src="https://aopwiki.org/system/dragonfly/production/2022/03/21/2gkpopb51s_Table_1._Sodium_channel_a_subunit.jpg" style="height:691px; width:940px" /></span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">This complex ontogeny of VGSCs confounds any simple linkage of VGSCs to adverse outcomes and is an uncertainty in the development of this AOP. Since brain development in both humans and rodents extends from early gestation through lactation, it is not currently possible to state which VGSC subtype, or subtypes, may be responsible for the AOs.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Ion channels, including VGSCs, are also expressed in oligodendrocytes, Schwann cells (Baker, 2002) and microglia (Hossain et al., 2017). The expression and function of VGSS in cells of the oligodendrocyte lineage follow a time and regional ontogeny. While present and active in the early stages of oligodendrocyte maturation, VGSC function decreases over developmental time and is absent in mature oligodendrocytes (Paez et al., 2009; Berret et al., 2017). Knockdown of VGSC in rat oligodendrocyte precursor cells (OPCs) leads to reduced myelination suggesting a function of VGSC for axon myelination (Berret et al., 2017).</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">The physiological and anatomical ontogeny of Schwann cells is well known (Jessen and Mirsky, 2005). VGSCs are present in Schwann cells including the tetrodotoxin sensitive and Na<sub>v</sub> 1.7 types (Ritche, 1992; Chiu, 1991; Baker, 2002) less is known about their developmental profile.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Microglia cells express several ion channels, including Cl<sup><span style="font-family:Symbol">-</span></sup>, K<sup>+</sup>, H<sup>+</sup> and Ca<sup>2+</sup> and VGSC that are involved in several cellular functions such as maintaining the membrane potential, cellular volume and intracellular ion concentrations. VGSCs are demonstrated, to be present both in rodents and human microglia. Different isoforms are present in primary microglia (Na<sub>v</sub> 1.1, 1.2, 1.3, 1.5, 1.6, 1.7, 1.8, 1.9, and 2.1 isoforms) compared to immortalised BV2 cells (Na<sub>v</sub> 1.2, 1.3, 1.4, 1.6, 1.8, 1.9, and 2.1 isoforms) (Jung et al., 2013; Black and Waxman, 2012; reviewed by Hossain et al., 2017). Presence of sodium channel isoforms in immortalised BV2 cells and primary microglia were detected by mRNA expression with standard PCR. BV2 cells express some sodium channel isoforms including Na<sub>v</sub> 1.2, 1.3, 1.4, 1.6, 1.8, 1.9, and 2.1 whereas primary microglia from 1–2-day-old mice express channel isoforms Na<sub>v</sub> 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7 1.8, 1.9, and 2.1. Primary microglia expressed higher levels of Na<sub>v</sub> 1.1. 1.2, 1.3, 1.6, 1.9, and 2.1 compared with BV2 cells.</span></span></p>
Not SpecifiedMaleNot SpecifiedFemaleNot SpecifiedAll life stagesNot SpecifiedNot Specified<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Baker MD, 2002. Electrophysiology of mammalian Schwann cells. Progress in Biophysics and Molecular Biology, 78(2–3), 83–103. <a href="https://doi.org/10.1016/S0079-6107(02)00007-X" style="color:blue; text-decoration:underline">https://doi.org/10.1016/S0079–6107(02)00007-X</a></span></span></span></p>
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<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black"><span style="background-color:white">Smith TJ and Soderlund DM, 1998. Action of the pyrethroid insecticide cypermethrin on rat brain IIa sodium channels expressed in xenopus oocytes. Neurotoxicology, 19(6), 823–832.</span></span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black"><span style="background-color:white">Smith TJ and Soderlund DM, 2001. Potent actions of the pyrethroid insecticides cismethrin and cypermethrin on rat tetrodotoxin-resistant peripheral nerve (SNS/PN3) sodium channels expressed in <em>Xenopus</em> oocytes. Pesticide Biochemistry and Physiology, 70(1), 52–61.</span> <a href="https://doi.org/10.1006/pest.2001.2538" style="color:blue; text-decoration:underline"><span style="background-color:white">https://doi.org/10.1006/pest.2001.2538</span></a></span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black"><span style="background-color:white">Smith TJ, Lee SH, Ingles PJ, Knipple DC and Soderlund DM, 1997. The L1014F point mutation in the house fly Vssc1 sodium channel confers knockdown resistance to pyrethroids. Insect Biochemistry and Molecular Biology, 27(10), 807–812.</span> <a href="https://doi.org/10.1016/S0965-1748(97)00065-9" style="color:blue; text-decoration:underline"><span style="background-color:white">https://doi.org/10.1016/S0965–1748(97)00065–9</span></a></span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black"><span style="background-color:white">Soderlund DM, Clark JM, Sheets LP, Mullin LS, Piccirillo VJ, Sargent D, </span>…<span style="background-color:white"> and Weiner ML, 2002. Mechanisms of pyrethroid neurotoxicity: implications for cumulative risk assessment. Toxicology, 171(1), 3–59.</span> <a href="https://doi.org/10.1016/S0300-483X(01)00569-8" style="color:blue; text-decoration:underline"><span style="background-color:white">https://doi.org/10.1016/S0300–483X(01)00569–8</span></a></span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black"><span style="background-color:white">Trainer VL, McPhee JC, Boutelet-Bochan H, Baker C, Scheuer T, Babin D, </span>…<span style="background-color:white"> and Catterall WA, 1997. High affinity binding of pyrethroids to the α subunit of brain sodium channels. Molecular Pharmacology, 51(4), 651–657.</span> <span style="background-color:white">doi: </span><a href="https://doi.org/10.1124/mol.51.4.651" style="color:blue; text-decoration:underline"><span style="background-color:white">https://doi.org/10.1124/mol.51.4.651</span></a></span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black"><span style="background-color:white">Vais H, Williamson MS, Devonshire AL and Usherwood PNR, 2001. The molecular interactions of pyrethroid insecticides with insect and mammalian sodium channels. Pest Management Science, 57(10), 877–888.</span> <a href="https://doi.org/10.1002/ps.392" style="color:blue; text-decoration:underline"><span style="background-color:white">https://doi.org/10.1002/ps.392</span></a></span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black"><span style="background-color:white">Vais H, Williamson MS, Goodson SJ, Devonshire AL, Warmke JW, Usherwood PN and Cohen CJ, 2000. Activation of <em>Drosophila</em> sodium channels promotes modification by deltamethrin: reductions in affinity caused by knock-down resistance mutations. Journal of General Physiology, 115(3), 305–318.</span> <span style="background-color:white">doi: </span><a href="https://doi.org/10.1085/jgp.115.3.305" style="color:blue; text-decoration:underline"><span style="background-color:white">https://doi.org/10.1085/jgp.115.3.305</span></a></span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black"><span style="background-color:white">Volpe JJ, Kinney HC, Jensen FE and Rosenberg PA, 2011. Reprint of ‘The developing oligodendrocyte: key cellular target in brain injury in the premature infant’. International Journal of Developmental Neuroscience, 29(6), 565–582. </span><a href="https://doi.org/10.1016/j.ijdevneu.2011.07.008" style="color:blue; text-decoration:underline"><span style="background-color:white"><span style="color:black">https://doi.org/10.1016/j.ijdevneu.2011.07.008</span></span></a></span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Wakeling EN, Neal AP and Atchison WD, 2012. Pyrethroids and their effects on ion channels. Pesticides—Advances in Chemical and Botanical Pesticides. Rijeka, Croatia: InTech, 39–66. <a href="http://dx.doi.org/10.5772/5033" style="color:blue; text-decoration:underline">http://dx.doi.org/10.5772/5033</a></span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black"><span style="background-color:white">Wang SY, Barile M and Wang GK, 2001. </span><span style="background-color:white">A phenylalanine residue at segment D3-S6 in </span>Na<sub>v</sub><span style="background-color:white">1.4 voltage-gated </span>Na<sup>+</sup><span style="background-color:white"> channels is critical for pyrethroid action. Molecular Pharmacology, 60(3), 620–628.</span></span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black"><span style="background-color:white">Westenbroek RE, Bischoff S, Fu Y, Maier SK, Catterall WA and Scheuer T, 2013. </span><span style="background-color:white">Localization of sodium channel subtypes in mouse ventricular myocytes using quantitative immunocytochemistry. Journal of Molecular and Cellular Cardiology, 64, 69–78. doi: 10.1016/j.yjmcc.2013.08.004</span></span></span></span></p>
2017-04-14T15:25:302023-11-10T03:36:24Disruption of sodium channel gating kineticsAltered kinetics of sodium channelCellular<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Action potentials are a temporary shift (from negative to positive) in the neuron’s membrane potential caused by ions flowing in and out of the neuron. During the resting state, before an action potential occurs, all the gated sodium and potassium channels are closed. These gated channels only open once when an action potential has been triggered. They are called ‘voltage-gated’ because they are open and close depending on the voltage difference across the cell membrane. VGSC have two gates (gate m and gate h), while the potassium channel only has one (gate n). Gate m (the activation gate) is normally closed and opens when the cell starts to get more positive. Gate h (the deactivation gate) is normally open, and swings shut when the cells gets too positive. Gate n is normally closed, but slowly opens when the cell is depolarised (very positive). VGSC exist in one of three states: Deactivated (closed) – at rest, channels are deactivated. The m gate is closed and does not let sodium ions through. Activated (open) – when a current pass through and changes the voltage difference across a membrane, the channel will activate and the m gate will open. Inactivated (closed) – as the neuron depolarises, the h gate swings shut and blocks sodium ions from entering the cell. Voltage-gated potassium channels are either open or closed.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Typically, activation and inactivation of VGSC occur quite rapidly. However, some compounds can bind to the VGSC and disrupt the kinetics of activation and inactivation. This typically slows the kinetics of those processes. Slowed VGSC activation leads to a decrease in peak Na<sup>+</sup> current measured throughout the cell as well as a delay in the time for the current to reach its peak. By slowing VGSC inactivation and deactivation leads to a prolonged VGSC open time. The longer channel open time results in more Na<sup>+</sup> entering the cell and this leads to hyperexcitability, membrane depolarisation, increase in firing rate and conduction block. A short prolongation of the channel inactivation kinetics causes repetitive firing of action potentials (repetitive discharge) as a small percentage of modified channels in the membrane can cause unmodified channels to activate, or open again. However, if the channel inactivation is sufficiently period, the membrane potential eventually becomes depolarised to the point at which generation of action potentials is not possible (depolarisation-dependent block). A small percentage of modified VGSCs can increase Na<sup>+</sup> current substantially (Narahashi, 1996), driving repetitive firing and depolarization-dependent conduction block.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">The modifications of the sodium channel gating have been studied using voltage- and patch-clamp experiments in models from many different invertebrate and vertebrate species, including mammals and even human cells (Ruigt et al., 1987; Soderlund et al., 2002), showing that the prolongation of the sodium current is mainly due to the reduced rate of closure of a fraction of sodium channels affected by pyrethroids. In neuroblastoma cell preparations, deltamethrin and other type II pyrethroids induced slow tail currents with a relatively rapid time constant. The rate at which sodium channels close during the pyrethroid-induced slow tail current depends not only on pyrethroid structure, but also on the time of exposure, temperature and membrane potential (Ruigt et al., 1987).</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">The voltage-clamp technique typically uses two microelectrodes, allowing control of the membrane potential and recording transmembrane currents flowing across the membrane of the entire cell that result from ion channel opening and closing (Guan et al., 2013).</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Patch clamp is a highly sensitive version of the voltage-clamp technique in which currents flowing through a single ion channel, or across the entire cell membrane ("whole cell" current) of an individual can be measured, depending on the configuration of the recording. Further, pharmacological interventions can be used such that the current flowing through only a single type of channel in the memberane can be measured. For example, by blocking potassium, calcium and chloride channels, the current flowing through voltage-gated sodium channels can be isolated. A single electrode serves both to measures voltage and pass current (Molleman, 2003). This technique allows measurement of the altered kinetics of a single channel, which can then be manifested by changes in the kinetics of the whole-cell current.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Ion channels are essential for the initiation and propagation of action potential in excitable cells from both vertebrate and invertebrate species. In neurons, ion channels are essential for chemical communication between cells, or synaptic transmission. Ion channels also function to maintain membrane potential and initiate and propagate electrical impulses. VGSC are therefore a target of natural and synthetic chemicals and disruption of the gate kinetics has been characterised in insects and mammalian cells (Soderlund et al., 2002).</span></span></p>
Not SpecifiedMaleNot SpecifiedFemaleNot SpecifiedAll life stagesNot SpecifiedNot Specified<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Guan B, Chen X and Zhang H, 2013. Two-electrode voltage clamp. Methods in Molecular Biology, 998, 79–89. doi: 10.1007/978-1-62703-351-0_6</span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Molleman A, 2003. Patch Clamping: An Introductory Guide to Patch Clamp Electrophysiology. John Wiley and Sons. </span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Narahashi T, 1996. Neuronal ion channels as the target sites of insecticides. Pharmacology and Toxicology, 79(1), 1–14. https://doi.org/10.1111/j.1600–0773.1996.tb00234.x</span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Ruigt GF, Neyt HC, Van der Zalm JM and Van den Bercken J, 1987. Increase of sodium current after pyrethroid insecticides in mouse neuroblastoma cells. Brain Research, 437(2), 309–322.</span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Soderlund DM, Clark JM, Sheets LP, Mullin LS, Piccirillo VJ, Sargen D, … and Weiner, ML, 2002. Mechanisms of pyrethroid neurotoxicity: implications for cumulative risk assessment. Toxicology, 171(1), 3–59. https://doi.org/10.1016/S0300–483X(01)00569–8</span></span></span></p>
2022-03-15T10:57:552022-09-13T20:44:31Disruption, action potentialDisruption in action potential generationCellular<p>An action potential is a fast, transitory and propagating change of the resting membrane potential. Neurons and muscle cells can generate an action potentials. The initial signal comes from other cells connecting to the neuron, and it causes positively charged ions to flow into the cell body. These ions pass through channels that open when a specific neurotransmitter binds to the channel, leading to opening. For example, when acetylcholine is released at the synapse between a nerve and muscle (called the neuromuscular junction) by a presynaptic neuron, it causes postsynaptic Na<sup>+</sup> channels to open. Na<sup>+</sup> enters the postsynaptic cell and causes the postsynaptic membrane to depolarise. This depolarisation is called an excitatory postsynaptic potential (EPSP) and makes the postsynaptic neuron more likely to fire an action potential. These incoming ions change the membrane potential closer to 0, a process known as depolarisation. When positive ions flow into the negative cell, the cell’s polarity decreases. If it gets positive enough, it can trigger the VGSC found in the axon, then the action potential will be sent.</p>
<p>This process lets positively charged sodium ions flow into the negatively charged axon and depolarise the surrounding axon. Once one channel opens and lets positive ions in, it sets the stage for the channels down the axon to perform the same thing in a domino-like process. This stage is known as depolarisation, the neuron becomes positively charged as the action potential passes through. When the inactivation gates of the sodium channels close, they stop the inward rush of positive ions. At the same time, the potassium channels open. There is much more potassium inside the cell compared with outside, so when these channels open, more potassium exits than enters. The cell therefore loses positively charged ions and returns back toward its resting state. This step is called repolarisation. As the action potential passes through, potassium channels stay open a little bit longer, and continue to let positive ions exit the neuron. This means that the cell temporarily hyperpolarises or gets even more negative than its resting state. As the potassium channels close, the sodium-potassium pump works to re-establish the resting state.</p>
<p>Sodium channel gating is a well regulated process that is critical to normal neuronal function, activation and propagation of the action potential. Shape, speed of conduction and fidelity in propagation of the action potential are essential to the timing, synchrony and efficacy of neuronal communication. Waveform, timing and fidelity of the axonal action potential can be modulated, which leads to changes in presynaptic neurotransmitter release. Action potential normally develops first in the initial segment of the axon. During axonal action potential initiation, the active depolarisation propagates both towards the soma (antidromic) and down the axon (orthodromic). The conduction velocity of the antidromic action potential may have a significant impact on dendritic backpropagation. This in turn will affect spike-timing dependent plasticity i.e. the synaptic plasticity sensitive to the timing of dendritic action potentials relative to incoming synaptic information. The orthodromic velocity will affect the degree of synchrony of arrival of information at different postsynaptic targets of the same axon. In neurons, voltage-gated sodium conductances play an essential role in action potential initiation and propagation. VGSC activate and inactivate within milliseconds. As the cell membrane is depolarised, sodium channels activate, resulting in the influx of sodium ions to further depolarise the membrane. This inward current produces the upstroke of the action potential. Along with the gating of potassium channels, sodium channel inactivation participates in the action potential downstroke. Although variations in many ion channels are likely to participate in the diversity of action potential waveforms observed in neurons, differences in sodium channel subunit composition, localisation and modulation may participate in shaping a neuron’s action potential. Sodium channel subunit composition at the axon initial segment contributes to the firing properties of neurons, particularly the characteristic maximum firing frequency of a particular cell class. Therefore, at nodes of Ranvier the sodium channel subunit composition may contribute to action potential propagation fidelity. Steady-state persistent sodium currents can contribute to excitability and to the shape of an action potential. These sodium channels are active near rest potential (−65 mV) and do not inactivate even with quite strong depolarisation. Therefore, these currents can participate in cellular excitability and in setting action potential threshold (Kress and Mennerick, 2009). Alterations in VGSCs can result in changes in membrane polarisation and propagation of neuronal action potentials. Changes in neuronal excitability in glutamatergic networks are described following treatment to deltamethrin and permethrin on neuronal activity in hippocampal neuronal cultures using patch clamp and microelectrode array (MEA) recordings (Meyer et al., 2008). Cao et al. (2011) demonstrated that VGSC responses of a neuronal network to pyrethroids with an increase of intracellular calcium concentration and these responses are secondary to activation of VGSCs. The effect of pyrethroids on neurotransmitters release and neuronal excitability in glutamatergic networks are described following treatment to deltamethrin and permethrin on neuronal activity in hippocampal neuronal cultures using patch clamp and microelectrode array (MEA) recordings (Meyer et al., 2008). The distinct abilities of pyrethroids to elevate BDNF mRNA expression are consistent with the demonstration of a range of pyrethroid efficacies in the stimulation of calcium influx. <em>In vivo</em>, deltamethrin has been reported to increase BDNF in the cortex and hippocampus (Imamura et al., 2006; Cao et al., 2011), and both deltamethrin and permethrin alter transcription profiles of activity-dependent genes in the cortex including c-fos, Egr1, and Camk1g (Harrill et al., 2008; Cao et al., 2011). Therefore, activity-dependent changes in gene transcription after pyrethroid exposure can occur both <em>in vitro</em> and <em>in vivo</em> (Cao et al., 2011; Pitzer et al., 2019; Zhang et al., 2018).</p>
<p>The action potential is a cycle of membrane depolarisation, hyperpolarisation and return to the resting value. To measure an action potential, the patch clamp or the intracellular recording (impale a sharp electrode into the cell cytosol) technique are generally used. For either, a glass-made microelectrode is sufficient to measure action potential. The measurement of Na+ ion concentration would not detect single action potentials but a change in bulk ion concentration over a longer time and that might depend mainly on the firing rate of the cells and the activity of Na+/K+-pumps. Patch clamp is the preferred technique for the qualification and quantification of the altered firing rate (Meyer et al., 2008). Neurotransmitter release can be evaluated in vivo using western blotting quantification or using microdialysis and analytical quantification.</p>
<p>Action potentials or nerve impulses are rapid and transient electrical activity that is propagated in the membrane of excitable such as neurons and muscle cells. Action potentials allow long-distance signalling in the nervous system. An action potential results from the sequential opening and closing of voltage-gated cation channels. First, opening of Na+ channels permits influx of Na+ ions for about 1 ms, causing a sudden large depolarisation of a segment of the membrane. The channel then closes and becomes unable to open (refractory) for several milliseconds, preventing further Na+ flow. Opening of K+ channels as the action potential reaches its peak permits efflux of K+ ions, which initially hyperpolarises the membrane. As these channels close, the membrane returns to its resting potential. The same basic mechanism is used by all neurons. Myelination produced by oligodendrocytes increases the velocity of impulse conduction (Lodish et al., 2000, ‘The Action Potential and Conduction of Electric Impulses’ in Molecular Cell Biology Section 21.2, New York )</p>
Not SpecifiedMaleNot SpecifiedFemaleNot SpecifiedAll life stagesHighHigh<p>Cao Z, Shafer TJ and Murray TF, 2011. Mechanisms of pyrethroid insecticide-induced stimulation of calcium influx in neocortical neurons, Journal of Pharmacology and Experimental Therapeutics, 336(1), 197–205. doi: https://doi.org/10.1124/jpet.110.171850</p>
<p>Harrill JA, Li Z, Wright FA, Radio NM, Mundy WR, Tornero-Velez R and Crofton KM, 2008. Transcriptional response of rat frontal cortex following acute in vivo exposure to the pyrethroid insecticides permethrin and deltamethrin. BMC Genomics, 9(1), 546. https://doi.org/10.1186/1471-2164-9-546</p>
<p>Imamura L, Yasuda M, Kuramitsu K, Hara D, Tabuchi A and Tsuda M, 2006. Deltamethrin, a pyrethroid insecticide, is a potent inducer for the activity-dependent gene expression of brain-derived neurotrophic factor in neurons. Journal of Pharmacology and Experimental Therapeutics, 316(1), 136–143. doi: 10.1124/jpet.105.092478</p>
<p>Kress GJ and Mennerick S, 2009. Action potential initiation and propagation: upstream influences on neurotransmission. Neuroscience, 158(1), 211–222.</p>
<p>Lodish H, Berk A, Zipursky SL, Matsudaira P, Baltimore D and Darnell J. Molecular Cell Biology. 4th edition. New York: W. H. Freeman; 2000. Section 21.2, The Action Potential and Conduction of Electric Impulses. Available from: https://www.ncbi.nlm.nih.gov/books/NBK21668/</p>
<p>Meyer DA, Carter JM, Johnstone AF and Shafer TJ, 2008. Pyrethroid modulation of spontaneous neuronal excitability and neurotransmission in hippocampal neurons in culture. Neurotoxicology, 29(2), 213– 225. doi: 10.1016/j.neuro.2007.11.005.</p>
<p>Pitzer EM, Sugimoto C, Gudelsky GA, Huff Adams CL, Williams MT and Vorhees CV, 2019. Deltamethrin exposure daily from postnatal day 3–20 in Sprague-Dawley rats causes long-term cognitive and behavioral deficits. Toxicological Sciences, 169(2), 511–523. https://doi.org/10.1093/toxsci/kfz067 Zhang C, Xu Q, Xiao X, Li W, Kang Q,</p>
<p>Zhang X, … and Li Y, 2018. Prenatal deltamethrin exposureinduced cognitive impairment in offspring is ameliorated by memantine through NMDAR/BDNF signaling in hippocampus, Frontiers in Neuroscience, 12, 615. https://doi.org/10.3389/fnins.2018.00615</p>
2022-03-31T06:45:062022-03-31T06:46:05Altered neurotransmission in developmentneurotrasmission in developmentCellular<p style="text-align:justify"><span style="font-size:11.0pt"><span style="font-family:"Times New Roman",serif">During axonal action potential initiation, the active depolarization propagates both towards the soma (antidromic) and down the axon (orthodromic). Because of very different passive and active membrane properties of the soma compared with the axon, the conduction velocity in the two directions is likely different in most cells. The conduction velocity of the antidromic action potential may have a significant impact on dendritic backpropagation. This in turn will affect spike-timing dependent plasticity, synaptic plasticity sensitive to the timing of dendritic action potentials relative to incoming synaptic information. The orthodromic velocity will affect the degree of synchrony of arrival of information at different postsynaptic targets of the same axon (Kress G.)</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">The arrival of the nerve impulse at the presynaptic terminal stimulates the release of neurotransmitter into the synaptic gap. The neuron is a secretory cell and the secretory product, the neurotransmitter, is released at the level of chemical synapses. </span></span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">Neurotransmitters synthesized by the neuron are stored in the presynaptic element, inside the </span><a href="https://www.sciencedirect.com/topics/neuroscience/synaptic-vesicle"><span style="font-family:"Times New Roman",serif">synaptic vesicles</span></a><span style="font-family:"Times New Roman",serif">. In the absence of presynaptic activity, the probability of a neurotransmitter being released in the synaptic cleft is very low. This probability increases strongly when the presynaptic element is depolarized by an action potential</span><span style="font-size:13.5pt"><span style="font-family:"Georgia",serif"><span style="color:#2e2e2e">. </span></span></span><span style="font-family:"Times New Roman",serif"> When an action potential reaches the axon terminal it depolarizes the membrane and opens VGSC. Sodium ions enter the cell, further depolarizing the presynaptic membrane. This depolarization causes voltage-gated Ca2+ channels to open. Calcium ions entering the cell initiate a signalling cascade that causes small membrane-bound vesicles, called synaptic vesicles, containing neurotransmitter molecules to fuse with the presynaptic membrane. Fusion of a vesicle with the presynaptic membrane causes neurotransmitter to be released into the synaptic cleft, the extracellular space between the presynaptic and postsynaptic membranes. The neurotransmitter diffuses across the synaptic cleft and binds to receptor proteins on the postsynaptic membrane. Once neurotransmission has occurred, the neurotransmitter is removed from the synaptic cleft and the postsynaptic membrane can “reset” and be ready to receive another signal. This can be accomplished in three ways: the neurotransmitter can diffuse away from the synaptic cleft, it can be degraded by enzymes in the synaptic cleft, or it can be recycled (sometimes called reuptake) by glia cells and the presynaptic neuron.</span></span></span></p>
<p style="text-align:justify"><span style="font-size:11pt"><span style="font-family:Calibri,sans-serif"><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif">In principle neurons can excrete excitatory and inhibitory neurotransmitters, which induces in the postsynaptic membrane either a depolarisation or hyperpolarisation, respectively. In consequence this can trigger or impede the generation of a new postsynaptic action potential. Neurons integrate the various excitatory and inhibitory signals they receive from the synapses with their presynaptic network, which leads to a net signalling result for their postsynaptic neurons. The development and function of these neurotransmissions can be disturbed by various mechanisms. </span></span></span></span><span style="font-size:11pt"><span style="font-family:Calibri,sans-serif"><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif">A useful indicative summary of potential effects of stressors on neurotransmission can be found, e.g. in the open textbook Foundations of Neuroscience by Casey Henley</span></span><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif"><span style="color:#00b050"> (</span></span></span><a href="https://openbooks.lib.msu.edu/neuroscience/chapter/drug-and-toxin-effects/" style="color:blue; text-decoration:underline"><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif">https://openbooks.lib.msu.edu/neuroscience/chapter/drug-and-toxin-effects/</span></span></a><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif"><span style="color:#00b050"> )</span></span></span></span></span></p>
<p style="text-align:justify"><span style="font-size:11pt"><span style="font-family:Calibri,sans-serif"><em><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif">“Drugs can alter neurotransmitter synthesis pathways, either increasing or decreasing the amount of neurotransmitter made in the terminal, affecting how much transmitter is released. An example of this is administration of L-DOPA, a dopamine precursor molecule that results in increased dopamine production; it is used as a treatment for Parkinson’s Disease. </span></span></em></span></span><span style="font-size:11pt"><span style="font-family:Calibri,sans-serif"><em><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif">Neurotransmitter packaging is another site of possible drug action. Reserpine, which has been used to treat high blood pressure, blocks the transport of the monoamine transmitters into vesicles by inhibiting the vesicular monoamine transporter (VMAT). This decreases the amount of neurotransmitter stores and the amount of neurotransmitter released in response to an action potential. </span></span></em></span></span><span style="font-size:11pt"><span style="font-family:Calibri,sans-serif"><em><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif">The neurotransmitter receptors are another critical location for drug and toxin action. Agonists mimic neurotransmitter effects, whereas antagonists block neurotransmitter effects. Muscimol, a component of some mushrooms, is an agonist for the ionotropic GABA receptor. Bicuculine, a component of some plants, is an antagonist to this receptor and blocks the action of GABA. Additionally, many chemicals are able to modulate receptors in either a positive or negative fashion. Alcohol binds to the GABA receptor and increases the time the receptor is open when GABA binds. </span></span></em></span></span><span style="font-size:11pt"><span style="font-family:Calibri,sans-serif"><em><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif">Finally, neurotransmitter degradation and reuptake can also be altered by drugs and toxins. Depending on the neurotransmitter, enzymes located in either the synapse or in the terminal are responsible for degradation of the transmitter, and these enzyme can be blocked by drugs. Organophosphates are found in many pesticides and prevent the action of acetylcholinesterase, the enzyme that breaks down acetylcholine in the synapse. This inhibition increases acetylcholine action on the postsynaptic neuron. Monoamine oxidase inhibitors (MAOIs) prevent monoamine oxidase from degrading the biogenic amine neurotransmitters. MAOIs have been used as antidepressants since they increase the amount of transmitter available. Additionally, drugs can prevent the reuptake of neurotransmitters into the presynaptic terminal. Cocaine blocks the dopamine transporter, which results in increased action of dopamine in the synapse. </span></span></em></span></span><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><em><span style="font-family:"Times New Roman",serif">Drugs and toxins can also affect neuron function by acting outside of the synapse. For example, some chemicals change voltage-gated ion channel dynamics. Veratridine, a compound found in plants from the lily family, prevents voltage-gated sodium channels from inactivating. Initially, this causes an increase in neurotransmitter release, but it can quickly lead to excitotoxicity.”</span></em> </span></span></p>
<p style="text-align:justify"><span style="font-size:11.0pt"><span style="font-family:"Times New Roman",serif">Neurotoxins acting on sodium channels have similar effects, at the steady state, on increase of Na influx, depolarization, Ca2 increase, and exocytosis. Depending on the toxin binding site on sodium channels, the release of neurotransmitter can be modified qualitatively and/or quantitatively (Messensini et al. 2003).</span></span></p>
<p style="text-align:justify"><span style="font-size:11.0pt"><span style="font-family:"Times New Roman",serif"><strong><img alt="" src="https://aopwiki.org/system/dragonfly/production/2022/05/16/6zegq2myxy_Mechanism_involved_in_synaptic_neurotransmitter_release.jpg" /></strong></span></span></p>
<p style="text-align:justify"><span style="font-size:11px">Figure 7. Mechanism involved in synaptic neurotransmitter release (Source: Encyclopedia Britannica) </span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">Neurotransmitter release can be measured by electrical effects of released neurotransmitter on postsynaptic membrane receptors and directly by biochemical assay. One way to estimate neurotransmitter release is to measure the postsynaptic response that it evokes. It is an indirect measure since it includes events following release, such as neurotransmitter diffusion from the pre- to the postsynaptic element, binding of neurotransmitter molecules to postsynaptic receptors and induction of the postsynaptic current. </span></span></span><span style="font-size:11pt"><span style="font-family:Calibri,sans-serif"><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif">Within scientific experiments often the influence of various biological modifiers (proteins, genes, chemicals) on the neurotransmission of inhibitory neurons and excitatory neurons is tested to understand the development, function and disturbance of neuronal networks. </span></span></span></span><span style="font-size:11pt"><span style="font-family:Calibri,sans-serif"><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif">This can be done, e.g. by methods that measure evoked inhibitory postsynaptic currents (eIPSCs) or evoked excitatory postsynaptic currents (eEPSCs) or micro postsynaptic currents (mPSCs). </span></span></span></span></p>
<p style="text-align:justify"><span style="font-size:11pt"><span style="font-family:Calibri,sans-serif"><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif">All these methods require to control spontaneous action potentials and pre-synaptic currents. This allows to use the patch clamp technique applying (evoking) pre-synaptic currents and measuring resulting post-synaptic currents (ePSCs) or measure just the mPSCs (thought to represent a response that is elicited by a single vesicle of transmitter under the condition of a pre-synaptically blocked action potential). </span></span></span></span><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">The electrophysiological technique used to measure such variables is the patch clamp (Neher & Marty, 1982) in the whole-cell recording mode, where the plasma membrane patch in the pipette is ruptured. The transverse hippocampal slice is widely used as an electrophysiological preparation to study synaptic plasticity. </span></span></span><span style="font-size:11pt"><span style="font-family:Calibri,sans-serif"><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif">The optimizations of the experimental settings for these measurements represent specific measurements for the quantitative relationship between evoking pre-synaptic current and resulting post-synaptic current. This is the immediate empirical evidence for this KE and associated KERs (KER3 in this AOP) in terms of essentially and, as such, can be used in the assessment of dose-concordance to empirically support the KER (KER3).</span></span></span></span></p>
<p style="text-align:justify"><span style="font-size:12pt"><span style="font-family:"Times New Roman",serif"><span style="font-size:10.0pt">Neurotransmitter release can also be examined using the genetically encoded synaptic transmission reporter synapto-pHluorin, which uses pH-sensitive mutants of GFP. The interiors of synaptic vesicles are acidified to a pH of approximately 5.7, an environment that keeps the pHluorins in an off state. When the vesicle fuses with the plasma membrane during exocytosis, the pH rises to extracellular levels, switching the pHluorin on and causing it to fluoresce (Figure 7.6). One of the advantages of using synapto-pHluorins is that the signal regenerates through multiple rounds of vesicle release and recycling, which permits vesicle recycling to be imaged in addition to synaptic transmission (Carter and Shieh, 2015).</span></span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">Neurotransmitter release can also be evaluated in vivo by western blotting quantification or by micro dialysis and analytical quantification.</span></span></span></p>
<p style="text-align:justify"><span style="font-size:12pt"><span style="font-family:"Times New Roman",serif"><span style="font-size:11.0pt">Patch Clamp</span></span></span></p>
<p style="text-align:justify"><span style="font-size:12pt"><span style="font-family:"Times New Roman",serif"><span style="font-size:10.0pt"><span style="background-color:white"><span style="color:#333333">The Patch-clamping is a versatile electrophysiological tool for understanding ion channel behaviour. Every cell expresses ion channels, but the most common cells to study with patch-clamp techniques include neurons, muscle fibres, cardiomyocytes, and oocytes overexpressing single ion channels.</span></span></span></span></span></p>
<p style="text-align:justify"><span style="font-size:12pt"><span style="font-family:"Times New Roman",serif"><span style="font-size:10.0pt">The patch-clamp technique involves a glass micropipette forming a tight gigaohm (GΩ) seal with the cell membrane. The micropipette contains a wire bathed in an electrolytic solution to conduct ions. The whole-cell technique involves rupturing a patch of membrane with mild suction to provide low-resistance electrical access, allowing control of transmembrane voltage. Alternatively, investigators can pull a patch of membrane away from the cell and evaluate currents through single channels via the inside-out or outside-out patch-c</span><span style="font-size:11.0pt">lamp technique.</span></span></span></p>
<p style="text-align:justify"><span style="font-size:12pt"><span style="font-family:"Times New Roman",serif"><span style="font-size:10.0pt">The analysis of single cell membranes can give information about how the cell membrane responds when changes to ionic strength, voltage potential, or cell type are changed. Patch clamp analysis is often studied in Petri dishes or well-plates to minimize manipulation and increase the chances of cell viability. However, planar electrical stimulation geometries can also be used for ease of use. Additionally, robotic targeting algorithms can be used to improve the rate of specified cell targeting.</span></span></span></p>
<p style="text-align:justify"><span style="font-size:12pt"><span style="font-family:"Times New Roman",serif"><span style="font-size:10.0pt">Voltage clumping</span></span></span></p>
<p style="text-align:justify"><span style="font-size:12pt"><span style="font-family:"Times New Roman",serif"><span style="font-size:10.0pt">Voltage clamping is performed by applying a sustained and consistent voltage to the sample to stimulate the membranes of excitable cells, initiating the ionic flow changes. These cells have a resting membrane potential and membrane resistance that must be measured and overcome before stimulation. Voltage clamping requires a voltage electrode for recording the transmembrane voltage and a current electrode for passing current through the membrane. A constant feedback loop of recording the membrane potential is generated, ensuring the cell remains at a constant potential. Once the desired potential has been stabilized, electrical potential measurements, as well as visual recordings are taken to determine the organelle functionality in the presence of various stimuli.</span></span></span></p>
<p style="text-align:justify"><span style="font-size:12pt"><span style="font-family:"Times New Roman",serif"><span style="font-size:10.0pt">Current clumping</span></span></span></p>
<p style="text-align:justify"><span style="font-size:12pt"><span style="font-family:"Times New Roman",serif"><span style="font-size:10.0pt">The current clamping technique applies a known current through the specimen and measures how the membrane potential changes. Unlike voltage clamping which seeks to maintain the membrane potential at a given value, current clamping uses a single micropipette to apply the desired current and record how the membrane potential changes in response. By administering repetitive current pulses, the membrane resistance can be calculated using Ohm’s law.</span></span></span></p>
<p style="text-align:justify"><span style="font-size:12pt"><span style="font-family:"Times New Roman",serif"><span style="font-size:10.0pt">Extracellular recording and microelectrodes array.</span></span></span></p>
<p><span style="font-size:11pt"><span style="background-color:whitesmoke"><span style="font-family:Calibri,sans-serif"><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif"><span style="color:black">Intracellular recordings are great for measurements of the ionic conditions in a single cell. A different set of measurements can look at changes in ion concentrations in the extracellular fluid or a group of neurons. Extracellular recordings show changes in the current or potential of several cells surrounding a microelectrode. Alterations to position and size of this electrode will change the nature of the measurement, depending on what properties are being investigated. By using two or more electrodes and a process called spike sorting, it is possible to work out the number of cells being recorded from and the activity occurring in each cell. Spike sorting uses computer algorithms to analyse the waveforms of the electrical activity from multiple electrodes and distinguish the activity of the individual neurons. Extracellular field potentials measure the electrical potential of a group of cells whose source is difficult to determine. The signals from these cells will overlap and the recording will be a sum of all of the electrical activity. These recordings are known as local field potentials.</span></span></span></span></span></span></p>
<p><span style="font-size:11pt"><span style="background-color:whitesmoke"><span style="font-family:Calibri,sans-serif"><span style="font-size:10.0pt"><span style="font-family:"Times New Roman",serif"><span style="color:black">Microelectrode arrays are chips that contain multiple electrodes through which neural signals can be recorded. They commonly have stimulating electrodes to deliver signals to a sample as well. The number of electrodes ranges from tens to thousands depending on the spatial resolution and amount of data required by the experiment. Different types of arrays can be used for a wide variety of in vitro and in vivo applications.</span></span></span></span></span></span></p>
<p><span style="font-size:11pt"><span style="font-family:"Calibri",sans-serif">Microelectrode array (MEA) recordings are also used to measure electrical activity in cultured neurons (Keefer et al., 2001; Gramowski et al., 2000; Gopal, 2003; Johnstone et al., 2010). MEAs can be applied in high-throughput platforms to facilitate screening of numerous chemical compounds (McConnell et al., 2012). Using selective agonists and antagonists of different classes of receptors their response can be evaluated in a quantitative manner (Novellino et al., 2011; Hogberg et al., 2011). The MEA allow examination of general network activity, bursting activity and network connectivity and using at least 16 measures it has been demonstrated to identify negative and positive control compounds, to identify concentrations at which network failure begins and to identify selective network perturbations versus non-specific cytotoxic effects when coupled with terminal cell death assays when using different test systems from different species (human, rats, mouse; Brown et al., 2016, 2017; Frank et al., 2017; Masjosthusmann et al., 2020; Vassallo et al., 2017).</span></span></p>
<p> </p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">The connections between neurons, and between neurons and downstream effector cells, occur at specialized cell junctions called synapses. Synapses can occur by direct electrical coupling between two cells, but chemical synapses, in which communication is via the release of a neurotransmitter, are more common and are involved in more complex information processing. The proteins involved in chemical synaptic transmission are much more numerous and diverse than those involved in electrical conduction In vertebrates, synaptic transmission usually travels in one direction, but ctenophore and cnidarian synapses are often bidirectional.</span></span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">We divide the synapse into (a) a presynaptic module, in which calcium signals are transduced into chemical secretions (known as excitation–secretion coupling); (b) a postsynaptic module (postsynaptic density), which comprises the proteins that support the specialized postsynaptic membrane and the signalling that goes on there; and (c) a module that determines the specific wiring diagram of neurons during development (axonogenesis). For the module “c”, during development, and after injury, axons must grow and find their correct synaptic targets. The proteins responsible for this targeting include secreted and membrane-bound signals and receptors that have not been studied in an evolutionary framework as well as for the other modules. Despite its apparent specialization for neuronal signalling, the excitation–secretion system in neurons comprises many ancient gene families. However, like the transduction module, these gene families are often used differently in the various animal lineages. The proteins involved in docking and in recycling are, for the most part, conserved across eukaryotes (Liebenskind et al. 2017).</span></span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">Several neurotransmitters have been found not only in animals, but also in plants and microorganisms. Thus, the presence of neurotransmitter compounds has been shown in organisms lacking a nervous system and even in unicellular organisms. Today, we have evidence that neurotransmitters, which participate in synaptic neurotransmission, are multifunctional substances participating in developmental processes of microorganisms, plants, and animals (Roschchina 2010).</span></span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">The neurotransmission wiring code, which includes Excitation–Secretion Coupling, Postsynaptic Density and Axonogenesis is present across multiple taxa and is representing a fundamental brain developmental process.</span></span></span></p>
HighMixedModerateDevelopment<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">Carter, M., & Shieh, J. C. (2015). Guide to research techniques in neuroscience. Academic Press.</span></span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">Encyclopædia Britannica, Inc. https://www.britannica.com/science/synapse, accessed on December 2020</span></span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">Liebeskind, B. J., Hofmann, H. A., Hillis, D. M., & Zakon, H. H. (2017). Evolution of animal neural systems, Annual review of ecology, evolution, and systematics, 48, 377-398, Annual Reviews, https://doi.org/10.1146/annurev-ecolsys-110316-023048. </span></span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">Massensini, A. R., Romano-Silva, M. A., & Gomez, M. V. (2003). </span><span style="font-family:"Times New Roman",serif">Sodium channel toxins and neurotransmitter release. Neurochemical research, 28(10), 1607-1611.</span></span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">Roshchina, V. V. (2010). Evolutionary considerations of neurotransmitters in microbial, plant, and animal cells. In Microbial endocrinology (pp. 17-52). Springer, New York, NY.</span></span></span></p>
<p style="text-align:justify"><span style="font-size:14px">Kress G.J. and Mennerick S. Action potential initiation and propagation: upstream influences on neurotransmission. <span style="font-family:Tahoma,sans-serif"><span style="font-family:"Times New Roman",serif">Neuroscience. 2009 January 12; 158(1): 211–222. doi:10.1016/j.neuroscience.2008.03.021)</span></span> </span></p>
2022-05-16T08:25:232022-05-18T11:39:32Hippocampal anatomy, Altered Hippocampal anatomy, Altered Tissue<p>The hippocampus is a major brain region located in the medial temporal lobe in humans and other mammals (West, 1990). Developmentally it is derived from neuronal and glial cells in the neural tube and differentiates in the proencephalon and telencephalon. The hippocampus is a cortical structure, but only contains 3-layers, distinct from the 6-layered neocortical structures. For this reason, it is known as archicortex or paleocortex meaning old cortex. Within humans, the structure is identified as early as fetal week 13 and matures rapidly until 2 to 3 years of age (Kier et al 1997), with continuing slow growth thereafter until adult ages (Utsunomiya et al., 1999). In rodents, the hippocampus begins to form in midgestation, with the CA fields forming in advance of the dentate gyrus. Dentate gyrus forms in late gestation with most of its development occurring in the first 2-3 postnatal weeks (Altman and Bayer, 1990a; 1990b).</p>
<p>The structure of the hippocampus has been divided into regions that include CA1 through CA4 and the dentate gyrus. The principal cell bodies of the CA field are pyramidal neurons, those of the dentate gyrus are granule cells. The dentate gyrus forms later in development than the CA fields of the hippocampus. These regions are generally found in all mammalian hippocampi.</p>
<p>The major input pathway to the hippocampus is from the layer 2 neurons of the entorhinal cortex to the dentate gyrus via the perforant path forming the first connection of the trisynaptic loop of the hippocampal circuit. Direct afferents from the dentate gyrus (mossy fibers) then synapse on CA3 pyramidal cells which in turn send their axons (Schaeffer Collaterals) to CA1 neurons to complete the trisynaptic circuit (Figure 1). From the CA fields information then passes through the subiculum entering the fiber pathways of the alveus, fimbria, and fornix and it routed to other areas of the brain (Amaral and Lavenex, 2006). Through the interconnectivity within the hippocampus and its connections to amygdala, septum and cortex, the hippocampus plays a pivotal role in several learning and memory processes, including spatial behaviors. The primary input pathway to the CA regions of the hippocampus is from the septum by way of the fornix and direct input from the amygdala. Reciprocal outputs from the hippocampus back to these regions and beyond also exist.</p>
<p><strong> </strong></p>
<p><strong>Trisynaptic Hippocampal Circuitry</strong></p>
<p><img alt="" 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" /></p>
<p style="text-align:justify"> </p>
<p>Data in support of this key event have been collected using a wide variety of standard biochemical, histological and anatomical methods (e.g., morphometrics, immunohistochemical staining, in situ hybridization and imaging procedures). Many of methods applied to reveal anatomical abnormalities are routine neurohistopathology procedures similar to those recommended in EPA and OECD developmental neurotoxicity guidelines (US EPA, 1998; OCED, 2007). Subtle cytoarchitectural features depend on more specialized birth dating procedures and staining techniques. It is essential to consider the timing of events during development for detection to occur, as well as the timing for detection (Hevner, 2007; Garman et al., 2001; Zgraggen et al., 2012). Similar techniques used in rodent stydies have been applied to postmortem tissue in humans. </p>
<p>In humans, structural neuroimaging techniques are used to assess hippocampal volume with an analysis technique known as voxel-based morphometry (VBM). Volume of brain regions is measured by drawing regions of interest (ROIs) on images from brain scans obtained from magnetic resonance imaging (MRI) or positron emission tomography (PET) scans and calculating the volume enclosed. (Mechelli et al., 2005). Similar imaging techniques can be applied in rodent models (Powell et al., 2009; Hasegawa et al., 2010; Pirko et al., 2005; Pirko and Johnson, 2008).</p>
<p>The hippocampus is generally similar in structure function across most mammalian species (West, 1990). The vast majority of information on the structure of the hippocampus is from mice, rats and primates including humans.</p>
UBERON:0000955brainHighMaleHighFemaleHighDuring brain developmentHighHighHigh<p>Altman J, Bayer SA. Migration and distribution of two populations of hippocampal granule cell precursors during the perinatal and postnatal periods. J Comp Neurol. 1990a Nov 15;301(3):365-81.</p>
<p>Altman J, Bayer SA. Prolonged sojourn of developing pyramidal cells in the intermediate zone of the hippocampus and their settling in the stratum pyramidale. J Comp Neurol. 1990b Nov 15;301(3):343-64.</p>
<p>Amaral D, Lavenex P (2006). "Ch 3. Hippocampal Neuroanatomy". In Andersen P, Morris R, Amaral D, Bliss T, O'Keefe J. The Hippocampus Book. Oxford University Press. ISBN 978-0-19-510027-3.</p>
<p>Garman RH, Fix AS, Jortner BS, Jensen KF, Hardisty JF, Claudio L, Ferenc S. Methods to identify and characterize developmental neurotoxicity for human health risk assessment. II: neuropathology. Environ Health Perspect. 2001 Mar;109 Suppl 1:93-100.</p>
<p>Hasegawa M, Kida I, Wada H. A volumetric analysis of the brain and hippocampus of rats rendered perinatal hypothyroid. Neurosci Lett. 2010 Aug 2;479(3):240-4.</p>
<p>Hevner RF. Layer-specific markers as probes for neuron type identity in human neocortex and malformations of cortical development. J Neuropathol Exp Neurol. 2007 66(2):101-9.</p>
<p>Kier, EL, Kim, JH, Fulbright, K, Bronen, RA. Embryology of the human fetal hippocampus: MR imaging, anatomy, and histology. AJNR Am J Neuroradiol: 1997, 18(3);525-32.</p>
<p>Mechelli A, Price C, Friston K, Ashburner J (2005) Voxel-Based Morphometry of the Human Brain: Methods and Applications. Curr Med Imaging Rev 1:105-113.</p>
<p>OECD. 2007. OECD guidelines for the testing of chemicals/ section 4: Health effects. Test no. 426: Developmental neurotoxicity study. http://www.oecd.org/dataoecd/20/52/37622194.</p>
<p>Pirko I, Fricke ST, Johnson AJ, Rodriguez M, Macura SI. Magnetic resonance imaging, microscopy, and spectroscopy of the central nervous system in experimental animals. NeuroRx. 2005 Apr;2(2):250-64.</p>
<p>Pirko I, Johnson AJ. Neuroimaging of demyelination and remyelination models. Curr Top Microbiol Immunol. 2008; 318:241-66.</p>
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2016-11-29T18:41:262022-05-20T05:45:05Altered function of the brainbrain functionOrgan2022-07-06T10:34:102022-07-06T10:34:10Impairment, Learning and memoryImpairment, Learning and memoryIndividual<p> </p>
<p>Learning can be defined as the process by which new information is acquired to establish knowledge by systematic study or by trial and error (Ono, 2009). Two types of learning are considered in neurobehavioral studies: a) associative learning and b) non-associative learning. Associative learning is based on making associations between different events. In associative learning, a subject learns the relationship among two different stimuli or between the stimulus and the subject’s behaviour. On the other hand, non-associative learning can be defined as an alteration in the behavioural response that occurs over time in response to a single type of stimulus. Habituation and sensitization are some examples of non-associative learning.</p>
<p>The memory formation requires acquisition, retention and retrieval of information in the brain, which is characterised by the non-conscious recall of information (Ono, 2009). There are three main categories of memory, including sensory memory, short-term or working memory (up to a few hours) and long-term memory (up to several days or even much longer).</p>
<p>Learning and memory depend upon the coordinated action of different brain regions and neurotransmitter systems constituting functionally integrated neural networks (D’Hooge and DeDeyn, 2001). Among the many brain areas engaged in the acquisition of, or retrieval of, a learned event, the hippocampal-based memory systems have received the most study. For example, the hippocampus has been shown to be critical for spatial-temporal memory, visio-spatial memory, verbal and narrative memory, and episodic and autobiographical memory (Burgess et al., 2000; Vorhees and Williams, 2014). However, there is substantial evidence that fundamental learning and memory functions are not mediated by the hippocampus alone but require a network that includes, in addition to the hippocampus, anterior thalamic nuclei, mammillary bodies cortex, cerebellum and basal ganglia (Aggleton and Brown, 1999; Doya, 2000; Mitchell et al., 2002, Toscano and Guilarte, 2005; Gilbert et al., 2006, 2016). Thus, damage to variety of brain structures can potentially lead to impairment of learning and memory. The main learning areas and pathways are similar in rodents and primates, including man (Eichenbaum, 2000; Stanton and Spear, 1990).While the prefrontal cortex and frontostriatal neuronal circuits have been identified as the primary sites of higher-order cognition in vertebrates, invertebrates utilize paired mushroom bodies, shown to contain ~300,000 neurons in honey bees (Menzel, 2012; Puig et al., 2014).</p>
<p>For the purposes of this KE (AO), impaired learning and memory is defined as an organism’s inability to establish new associative or non-associative relationships, or sensory, short-term or long-term memories which can be measured using different behavioural tests described below.</p>
<p><strong>In laboratory animals:</strong> in rodents, a variety of tests of learning and memory have been used to probe the integrity of hippocampal function. These include tests of spatial learning like the radial arm maze (RAM), the Barnes maze, <span style="color:#3498db">Hebb-Williams maze</span>, passive avoidance and Spontaneous alternation and most commonly, the Morris water maze (MWM). Test of novelty such as novel object recognition, and fear based context learning are also sensitive to hippocampal disruption. Finally, trace fear conditioning which incorporates a temporal component upon traditional amygdala-based fear learning engages the hippocampus. A brief description of these tasks follows.</p>
<p>1) RAM, Barnes, MWM, <span style="color:#3498db">Hebb-Williams maze </span>are examples of spatial tasks, animals are required to learn the location of a food reward (RAM); an escape hole to enter a preferred dark tunnel from a brightly lit open field area (Barnes maze), or a hidden platform submerged below the surface of the water in a large tank of water (MWM) (Vorhees and Williams, 2014). The <span style="color:#3498db">Hebb-Williams maze measures an animal’s problem solving abilities by providing no spatial cues to find the target (Pritchett & Mulder, 2004).</span></p>
<p>2) Novel Object recognition. This is a simpler task that can be used to probe recognition memory. Two objects are presented to animal in an open field on trial 1, and these are explored. On trial 2, one object is replaced with a novel object and time spent interacting with the novel object is taken evidence of memory retention – I have seen one of these objects before, but not this one (Cohen and Stackman, 2015).</p>
<p>3) Contextual Fear conditioning is a hippocampal based learning task in which animals are placed in a novel environment and allowed to explore for several minutes before delivery of an aversive stimulus, typically a mild foot shock. Upon reintroduction to this same environment in the future (typically 24-48 hours after original training), animals will limit their exploration, the context of this chamber being associated with an aversive event. The degree of suppression of activity after training is taken as evidence of retention, i.e., memory (Curzon et al., 2009).</p>
<p>4) Trace fear conditioning. Standard fear conditioning paradigms require animals to make an association between a neutral conditioning stimulus (CS, a light or a tone) and an aversive stimulus (US, a footshock). The unconditioned response (CR) that is elicited upon delivery of the footshock US is freezing behavior. With repetition of CS/US delivery, the previously neutral stimulus comes to elicit the freezing response. This type of learning is dependent on the amygdala, a brain region associated with, but distinct from the hippocampus. Introducing a brief delay between presentation of the neutral CS and the aversive US, a trace period, requires the engagement of the amygdala and the hippocampus (Shors et al., 2001).</p>
<p><span style="color:#3498db">5) Operant Responding. Performance on operant responding reflects the cortex’ ability to organize processes (Rabin et al., 2002). </span></p>
<p><strong>In humans:</strong> A variety of standardized learning and memory tests have been developed for human neuropsychological testing, including children (Rohlman et al., 2008). These include episodic autobiographical memory, perceptual motor tests, short and long term memory tests, working memory tasks, word pair recognition memory; object location recognition memory. Some have been incorporated in general tests of intelligence (IQ) such as the Wechsler Adult Intelligence Scale (WAIS) and the Wechsler. Modifications have been made and norms developed for incorporating of tests of learning and memory in children. Examples of some of these tests include:</p>
<p>1) Rey Osterieth Complex Figure test (RCFT) which probes a variety of functions including as visuospatial abilities, memory, attention, planning, and working memory (Shin et al., 2006).</p>
<p>2) Children’s Auditory Verbal Learning Test (CAVLT) is a free recall of presented word lists that yields measures of Immediate Memory Span, Level of Learning, Immediate Recall, Delayed Recall, Recognition Accuracy, and Total Intrusions. (Lezak 1994; Talley, 1986).</p>
<p>3) Continuous Visual Memory Test (CVMT) measures visual learning and memory. It is a free recall of presented pictures/objects rather than words but that yields similar measures of Immediate Memory Span, Level of Learning, Immediate Recall, Delayed Recall, Recognition Accuracy, and Total Intrusions. (Lezak, 1984; 1994).</p>
<p>4) Story Recall from Wechsler Memory Scale (WMS) Logical Memory Test Battery, a standardized neurospychological test designed to measure memory functions (Lezak, 1994; Talley, 1986).</p>
<p>5) Autobiographical memory (AM) is the recollection of specific personal events in a multifaceted higher order cognitive process. It includes episodic memory- remembering of past events specific in time and place, in contrast to semantic autobiographical memory is the recollection of personal facts, traits, and general knowledge. Episodic AM is associated with greater activation of the hippocampus and a later and more gradual developmental trajectory. Absence of episodic memory in early life (infantile amnesia) is thought to reflect immature hippocampal function (Herold et al., 2015; Fivush, 2011).</p>
<p>6) Staged Autobiographical Memory Task. In this version of the AM test, children participate in a staged event involving a tour of the hospital, perform a series of tasks (counting footprints in the hall, identifying objects in wall display, buy lunch, watched a video). It is designed to contain unique event happenings, place, time, visual/sensory/perceptual details. Four to five months later, interviews are conducted using Children’s Autobiographical Interview and scored according to standardized scheme (Willoughby et al., 2014).</p>
<p><span style="color:#3498db">7) Attentional set-shifting (ATSET) task. Measures the ability to relearn cues over various schedules of reinforcement (Heisler et al., 2015).</span></p>
<p>8. Comprehensive developmental inventory for infants and toddlers (CDIIT). The CDIIT was designed and standardized in 1996, and it measures the global, cognitive, language, motor, gross motor, fine motor, social, self-help and behavioral developmental status of children from 3 to 71 months old (Wang et al., 1998).</p>
<p><strong>In Honey Bees:</strong> For over 50 years an assay for evaluating olfactory conditioning of the proboscis extension reflex (PER) has been used as a reliable method for evaluating appetitive learning and memory in honey bees (Guirfa and Sandoz, 2012; LaLone et al., 2017). These experiments pair a conditioned stimulus (e.g., an odor) with an unconditioned stimulus (e.g., sucrose) provided immediately afterward, which elicits the proboscis extension (Menzel, 2012). After conditioning, the odor alone will lead to the conditioned PER. This methodology has aided in the elucidation of five types of olfactory memory phases in honey bee, which include early short-term memory, late short-term memory, mid-term memory, early long-term memory, and late long-term memory (Guirfa and Sandoz, 2012). These phases are dependent on the type of conditioned stimulus, the intensity of the unconditioned stimulus, the number of conditioning trials, and the time between trials. Where formation of short-term memory occurs minutes after conditioning and decays within minutes, memory consolidation or stabilization of a memory trace after initial acquisition leads to mid-term memory, which lasts 1 d and is characterized by activity of the cAMP-dependent PKA (Guirfa and Sandoz, 2012). Multiple conditioning trials increase the duration of the memory after learning and coincide with increased Ca2+-calmodulin-dependent PKC activity (Guirfa and Sandoz, 2012). Early long-term memory, where a conditioned response can be evoked days to weeks after conditioning requires translation of existing mRNA, whereas late long-term memory requires de novo gene transcription and can last for weeks (Guirfa andSandoz, 2012)."</p>
<p>Basic forms of learning behavior such as habituation have been found in many taxa from worms to humans (Alexander, 1990). More complex cognitive processes such as executive function likely reside only in higher mammalian species such as non-human primates and humans. Recently, larval zebrafish has also been suggested as a model for the study of learning and memory (Roberts et al., 2013).</p>
<p><span style="color:#3498db"><strong>Life stage applicability: </strong>This key event is applicable to various life stages such as during brain development and maturity (Hladik & Tapio, 2016). </span></p>
<p><span style="color:#3498db"><strong>Sex applicability:</strong> This key event is not sex specific (Cekanaviciute et al., 2018), although sex-dependent cognitive outcomes have been recently ; Parihar et al., 2020). </span></p>
<p><span style="color:#3498db"><strong>Evidence for perturbation by a prototypic stressor: </strong>Current literature provides ample evidence of impaired learning and memory being induced by ionizing radiation (Cekanaviciute et al., 2018; Hladik & Tapio, 2016). </span></p>
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2016-11-29T18:41:242023-06-26T12:44:45169bfc32-cee6-4cdf-ace4-d94aa23b43650ba805ac-676b-442b-85b3-adf93a790b9c<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">VGSCs are critical in generation and conduction of electrical signals in multiple excitable tissues. Natural and synthetic toxins are known to interact with VGSC by altering the gate kinetic of the channel by slowing the activation and deactivation rate of the VGSC and shift to a more hyperpolarised potentials the membrane potential at which the VGSC activate.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">The detailed mechanism of voltage sensing and voltage-dependent activation of the voltage sensor of sodium channels through a series of resting and activated states is known at the atomic level.</span></span></p>
<p><span style="font-size:11.0pt"><span style="font-family:"Calibri",sans-serif">There is evidence supporting that the binding of pyrethroids to VGSC (Trainer et al., 1997; O’Reilly et al., 2006) induces disruption of the sodium channel gate kinetics (Meyer et al., 2008; Soderlund et al., 2002).</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">It is well known that ion channels are integral membrane proteins that are critical for the execution of action potential and therefore for neuronal function and activation. Action potentials are the electrical impulses that travel along the axons of neurons and result from the movement of Na<sup>+</sup> and potassium (K<sup>+</sup>) ions across the membrane. Binding of excitatory neurotransmitters to their receptors opens cation-permeable ion channels causing the membrane to depolarise or become more positive. This depolarisation activates (opens) VGSCs allowing Na<sup>+</sup> to enter the neuron further depolarising the membrane. This increase in membrane permeability to Na<sup>+</sup> is responsible for the rising phase of the action potential, eventually causing the membrane polarity to reverse (overshoot phase). The falling phase of the action potential is caused by the inactivation of the VGSCs and the opening of voltage-gated potassium channels allowing K<sup>+</sup> to leave the cell. The efflux of K<sup>+</sup> ions results in hyperpolarisation (undershoot phase) of the membrane. Ultimately the voltage-gated K<sup>+</sup> channels close and the membrane potential returns to its resting state. Type I and II pyrethroids cause stimulus dependent membrane depolarisation and conduction block.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">It is therefore biologically plausible that binding of a chemical substance to a VGSC leads sodium channels to open at more hyperpolarised potentials and kept open longer (disruption of channel kinetic), allowing more sodium ions to cross and depolarise the neuronal membrane (Shafer et al., 2005)</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Expression of VGSC are spatially and temporally dependent; however, it is biologically plausible that also in developing brain pyrethroids would bind to VGSC isoforms and disrupt the channel gating kinetic (Shafer et al., 2005; Soderlund et al., 2002).</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Pyrethroids bind on the sodium channel α-subunit and affect nervous system function by altering their normal gating kinetics. Due to the extreme lipophilicity and the modest potency of pyrethroids radioligand, initial studies attempting to label the binding site were unsuccessful. The subsequent use of more potent radioligands were able to demonstrate high affinity saturable binding to brain sodium channels. However, the high lipophilicity of pyrethroids is still a limitation for the sensitivity of the assay and the identification of a single binding site on any given sodium channel and its mediated action (Soderlund et al., 2002; Trainer et al., 1997).</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">In hippocampal cell cultures from rat postnatal day 2–4 pups, patch clamp preparations of isolated neurons showed that deltamethrin alter the VGSC kinetic and inhibits neuronal activity in glutamatergic networks of hippocampal neurons in a potent and concentration-dependent manner (Meyer et al., 2008). Indeed, the actions of DLM are consistent with a decrease amplitude and number of spikes elicited using the current pulse (Meyer et al., 2008). <em>In vitro</em> exposure to pyrethroids (the type I permethrin and the type II deltamethrin) has been shown to differently disrupt sodium channel gate kinetics (Meyer et al., 2008) on hippocampal cultures from postnatal day 2–4 pups. This <em>in vitro</em> model was considered appropriate to explore effect on the developing brain. Cells were used for electrophysiological recording (patch clamp) 8–12 days <em>in vitro</em> (DIV) and hippocampal neurons isolated from early postnatal rodents form spontaneously active networks of interconnected neurons in which both glutamate and GABAergic neurotransmission occurs. Deltamethrin decreases neuronal excitability as measured by the rate of sEPSC activity at the concentration of 0.1 µM. At this concentration decrease in sEPSC interevent interval was rapid, occurring within 1–3 minutes of exposure and persistent, lasting throughout the exposure period (9 minutes). The effect on sEPSC frequency was concentration-dependent between 0.01 and 10 µM with an EC50 of 0.037 µM. There was no effect on the sEPSC amplitude at any tested concentration and this was consistent with previous data (Meyer and Shafer 2006), indicating that the effect does not include actions on postsynaptic glutamate receptors (Meyer et al., 2008).</span></span></p>
<p style="text-align:justify"> </p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">DOSE CONCORDANCE</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Although no evidence is available for the prototype stressor used in this AOP, deltamethrin, on the binding to VGSC, there is indirect evidence measuring the relationship between the MIE and the disruption of the VGSC gate kinetics. At concentration between 0.01 to 1 <span style="font-family:Symbol">m</span>M, deltamethrin has been shown to differently disrupt sodium channel gate kinetics (Meyer et al., 2008) on hippocampal cultures from postnatal day 2–4 pups. The effect was measured using the restricted patch clamp methodology and the results indicated that the observed change was concentration-dependent on the sEPSC without affecting the sEPSC amplitude and therefore excluding a postsynaptic excitatory mediated effect.</span></span></p>
<p style="text-align:justify"> </p>
<p style="text-align:justify"><span style="font-size:14px">TIME CONCORDANCE</span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Changes in the VGSC kinetics are evident immediately following exposure in vitro to deltamethrin and recorded up to 9 minutes (Mayer et al., 2008)</span></span></p>
<p style="text-align:justify"> </p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">The fact that binding of pyrethroids to VGSCs results in altered sodium channel gate kinetics is well accepted and supported by some evidence. However, some minor uncertainties can be detected as reported below.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Uncertainties in the overall knowledge remain as the sodium channels’ ontogeny is a complex process. Since brain development in both humans and rodents extends from early gestation through lactation it is not possible to state with certainty which isoform of the sodium channels’ α subunits is preferentially affected by deltamethrin.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">For <em>in vitro</em> methodologies, there is still a lack of knowledge on stability of deltamethrin in the medium and the partitioning of this compound with plastic, lipid and protein. Indeed, the high lipophilicity of pyrethroids is still a limitation for the sensitivity of the assays and for the identification of a single binding site on any given sodium channel and its mediated action this may affect the sensitivity of the assays (Ruigt et al., 1987). Also, the metabolic competence of the test systems used in various assays is unknown.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Moreover, the study from Meyer et al. (2008) is an indirect measurement of the interaction between the prototype stressor, deltamethrin and VGSCs. Also, the exact temperature at which the patch clamp recording was made is uncertain (in the publication it is stated at room temperature) and it is well documented that pyrethroids effects on VGSCs are negatively temperature dependent (reviewed in Narahashi, 2000). Finally, Meyer and colleagues used hippocampal cell culture from rats PND 2–4 which were not characterised and did not contain microglia or oligodendrocyte precursors cells, therefore there are still uncertainties in the knowledge of the interaction between pyrethroids and microglia or oligodendrocytes precursor VGSC.</span></span></p>
<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Some inconsistencies can be observed in experimental studies. They are associated with the electrophysiological technique used to study ionic currents in individual isolated living cells, tissue sections or patches of cells. The solution used in the bath can be similar to cytoplasm composition or completely different, they can be changed by adding ions or drugs to study the ion channels under different conditions. In the study of Meyer et al. (2008) different effects, i.e. burst duration, were observed for permethrin (type I) and deltamethrin (type II) and it was not clear if this represents a true difference in the mode of action between type I and type II pyrethroids or simply a difference between the two compounds. This could only be determined by the examination of additional chemicals.</span></span></p>
Not SpecifiedMaleNot SpecifiedFemaleNot SpecifiedAll life stagesNot SpecifiedNot Specified<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Chahine M (ed.), 2018. Voltage-gated Sodium Channels: Structure, Function and Channelopathies. Vol. 246. Springer.</span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Meisler MH, Kearney J, Ottman R and Escayg A, 2001. Identification of epilepsy genes in human and mouse. Annual Review of Genetics, 35(1), 567–588.</span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Meyer DA and Shafer TJ, 2006. Permethrin, but not deltamethrin, increases spontaneous glutamate release from hippocampal neurons in culture. Neurotoxicology, 27, 594–603.</span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Meyer DA, Carter JM, Johnstone AF and Shafer TJ, 2008. Pyrethroid modulation of spontaneous neuronal excitability and neurotransmission in hippocampal neurons in culture. Neurotoxicology, 29(2), 213–225. doi: 10.1016/j.neuro.2007.11.005.</span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Narahashi T, 2000. Neuroreceptors and ion channels as the basis for drug action: past, present, and future. J Pharmacol Exp Ther, 294, 1–26.</span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">O'Reilly AO, Khambay BP, Williamson MS, Field LM, Wallace BA and Davies TG, 2006. Modelling insecticide-binding sites in the voltage-gated sodium channel. Biochemical Journal, 396(2), 255–263.</span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Planells-Cases R, Caprini M, Zhang J, Rockenstein EM, Rivera RR, Murre C, … and Montal M, 2000. Neuronal death and perinatal lethality in voltage-gated sodium channel αII-deficient mice. Biophysical Journal, 78(6), 2878–2891.</span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Shafer TJ, Meyer DA and Crofton KM, 2005. Developmental neurotoxicity of pyrethroid insecticides: critical review and future research needs. Environmental Health Perspectives, 113(2), 123–136. https://doi.org/10.1289/ehp.7254.</span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Soderlund DM, Clark JM, Sheets LP, Mullin LS, Piccirillo VJ, Sargent D, … and Weiner ML, 2002. Mechanisms of pyrethroid neurotoxicity: implications for cumulative risk assessment. Toxicology, 171(1), 3–59. <a href="https://doi.org/10.1016/S0300-483X(01)00569-8" style="color:blue; text-decoration:underline">https://doi.org/10.1016/S0300–483X(01)00569–8</a></span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black"><span style="background-color:white">Trainer VL, McPhee JC, Boutelet-Bochan H, Baker C, Scheuer T, Babin D, </span>…<span style="background-color:white"> and Catterall WA, 1997. High affinity binding of pyrethroids to the α subunit of brain sodium channels. Molecular Pharmacology, 51(4), 651–657.</span> <span style="background-color:white">doi: </span><a href="https://doi.org/10.1124/mol.51.4.651" style="color:blue; text-decoration:underline"><span style="background-color:white">https://doi.org/10.1124/mol.51.4.651</span></a></span></span></span></p>
<p style="margin-left:19px; text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif"><span style="color:black">Wakeling EN, Neal AP and Atchison WD, 2012. Pyrethroids and their effects on ion channels. Pesticides—Advances in Chemical and Botanical Pesticides. Rijeka, Croatia: InTech, pp. 39–66. <a href="http://dx.doi.org/10.5772/50330" style="color:blue; text-decoration:underline">http://dx.doi.org/10.5772/50330</a></span></span></span></p>
2022-03-21T08:56:452022-03-31T06:51:510ba805ac-676b-442b-85b3-adf93a790b9c935308b9-c7a3-402c-b8fb-c562c53c613e<p>Long lasting modification of VGSC increases the channel opening time. The direct consequence is a more hyperpolarised potential. This underlies the disruption of neuronal activity with changes in the ions intracellular concentrations and neuronal excitability. Depending on the time the channel is left open, the disruption of the action potential is getting qualitative different and this difference is measurable in electrophysiological recording of the action potential. Limited opening will lead to repetitive firing while following prolonged opening the membrane potential ultimately becomes depolarised to the point at which generation of an action potential is not possible (depolarisation-dependent block) (Shafer et al., 2005).</p>
<p>The falling phase of the action potential caused by the inactivation of the VGSCs and the opening of voltage-gated potassium channels allowing K<sup>+</sup> to leave the cell. The efflux of K<sup>+</sup> ions results in hyperpolarisation (undershoot phase) of the membrane. Ultimately the voltage-gated K<sup>+</sup> channels close and the membrane potential returns to its resting state. It is therefore biologically plausible that changing the dynamic of VGSCs leads to a series of complex cellular events resulting in alteration of the firing rate as a final consequence. Type II pyrethroids cause stimulus dependent membrane depolarisation and conduction block. Expression of VGSC are spatial and temporal dependent; however, it is biological plausible that also in developing brain pyrethroids would bind to VGSC isoforms and disrupt the channel gating kinetic (Shafer et al., 2005; Soderlund et al., 2002).</p>
<p>Effect on the neuronal electrical activity using the type II pyrethroid deltamethrin is reported in Meyer et al. (2008) when using hippocampal cultures from postnatal day 2–4 pups. The electrical changes indicate neuron depolarisation and conduction block consequent to disruption of action potential generation with a dose-dependent inhibition of spontaneous glutamate release from hippocampal neurons. Deltamethrin inhibits spontaneous glutamate release from hippocampal neurons as measured by a decrease in sEPSC frequency during bursting release activity (Meyer et al., 2008). The effect is considered presynaptic because the decrease in sEPSC frequency following treatment with deltamethrin was not accompanied by changes in amplitude (Meyer et al., 2008). These data support the fact that deltamethrin decreases neuronal excitation by inhibition of the firing rate (inhibition of the spontaneous spiking activity) and the subsequent release of glutamate from the synapse.</p>
<p>Alterations of calcium dynamics are also reported for pyrethroids (Soderlund et al., 2002; Cao et al., 2011). Extracellular calcium, rather than calcium release from the intracellular calcium stores, is the likely source for pyrethroid-induced elevation of calcium in neocortical neurons (Cao et al., 2011). The same paper demonstrates that L-type VGCCs, NMDA receptors, and the sodium/calcium exchanger accounted for most pyrethroid-induced calcium entry. TTX completely abolished pyrethroid-induced calcium entry, indicating that these pathways were activated as a result of pyrethroid actions on VGSCs. For L-type VGCCs, activation by deltamethrin is likely to have been secondary to depolarisation of the cell membrane as a result of VGSC activation. Although it was not measured, it is likely that the depolarisation and calcium entry resulted in glutamate release, which then activated NMDA receptors, resulting in additional calcium entry. Finally, sodium entry through VGSC may have caused sodium loading of the neurons, which can result in a reversal of sodium/calcium exchange, which accounts for the contribution of this component to pyrethroid-induced calcium entry (Cao et al., 2011).</p>
<p><em>Dose Concordance</em></p>
<p>Changes in VGSC kinetic and disruption of the action potential are reported <em>in vitro</em> at concentration between 0.01 to 1 mM, in hippocampal or neocortical neurons from postnatal day 2–4 pups (Meyer et al., 2008; Cao et al., 2011).</p>
<p><em>Temporal Concordance</em></p>
<p>The two KEs were observed immediately following exposure to deltamethrin when measured <em>in vitro</em> up to 800 seconds recording in Cao et al. (2007) and immediately following exposure to deltamethrin when measured <em>in vitro</em> up to 9 minutes recording in Mayer et al., 2008.</p>
<p>The mechanistic understanding of the generation of membrane potentials, based on Na, K, Cl and Ca ions is broadly accepted and extensive documentation is also available. However, some uncertainties can be detected. The uncertainties and inconsistencies detected in the Meyer et al., 2008 are also applicable for this KER.</p>
<p>The events investigated by Cao et al. (2011) e.g. depolarisation and calcium entry, glutamate release, activation of NMDA receptors and additional calcium entry, were not directly measured in the study. Moreover, as reported also for VGSCs, the action of pyrethroids on calcium channel is temperature dependent and may have an impact on the deltamethrin-induced calcium influx in neocortical neurons. in the study from Cao et al. (2011) the temperature at which the experiment was carried out is not reported. 9 out of 11 pyrethroids tested were able to produce a concentration-dependent elevation in intracellular calcium concentration in neocortical neurons which occurred secondary to activation of VGSCs. The nine pyrethroids that stimulated calcium influx displayed distinct efficacies. The rank order of efficacy for calcium influx was similar to that for sodium influx (Cao et al., 2009) with the exception of S-bioallethrin, which is the least efficacious compound on calcium influx. Deltamethrin, the prototype stressor for this AOP, is in position 6 (out of 9) in terms of potency.</p>
<p>It should be further noted that other ionic channels may have an impact on the action potential generation and in this regard the knowledge is limited.</p>
<p>Also, in this case, some inconsistencies can be observed in experimental studies. They can be associated with the electrophysiological technique used to study ionic currents in individual isolated living cells, tissue sections or patches of cells. The solution used in the bath can be similar to cytoplasm composition or completely different, they can be changed by adding ions or drugs to study the ion channels under different conditions.</p>
Not SpecifiedMaleNot SpecifiedFemaleNot SpecifiedAll life stagesHighHigh<p>Cao Z, George J, Baden DG and Murray TF, 2007. Brevetoxin-induced phosphorylation of Pyk2 and Src in murine neocortical neurons involves distinct signaling pathways. Brain Res 1184:17–27.</p>
<p>Cao Z, Shafer TJ and Murray TF, 2009. Influence of pyrethroid insecticides on sodium and calcium influx in neocortical neurons. Toxicologist, 108, 443.</p>
<p>Cao Z, Shafer TJ and Murray TF, 2011. Mechanisms of pyrethroid insecticide-induced stimulation of calcium influx in neocortical neurons, Journal of Pharmacology and Experimental Therapeutics, 336 (1), 197–205. doi: https://doi.org/10.1124/jpet.110.171850</p>
<p>Meyer DA, Carter JM, Johnstone AF and Shafer TJ, 2008. Pyrethroid modulation of spontaneous neuronal excitability and neurotransmission in hippocampal neurons in culture. Neurotoxicology, 29(2), 213–225. doi: 10.1016/j.neuro.2007.11.005</p>
<p>Shafer TJ, Meyer DA and Crofton KM, 2005. Developmental neurotoxicity of pyrethroid insecticides: critical review and future research needs. Environmental Health Perspectives, 113(2), 123–136. <a href="https://doi.org/10.1289/ehp.7254">https://doi.org/10.1289/ehp.7254</a></p>
<p>Soderlund DM, Clark JM, Sheets LP, Mullin LS, Piccirillo VJ, Sargent D, … and Weiner ML, 2002. Mechanisms of pyrethroid neurotoxicity: implications for cumulative risk assessment. Toxicology, 171(1), 3–59. https://doi.org/10.1016/S0300–483X(01)00569–8.</p>
2022-03-31T06:53:002022-03-31T06:54:57Binding to voltage gate sodium channels during development leads to cognitive impairment Binding to VGSC during development leads to cognitive impairmentUnder development: Not open for comment. Do not citeUnder DevelopmentIncluded in OECD Work Plan1.91<p style="text-align:justify"><span style="font-size:10pt"><span style="font-family:Tahoma,sans-serif">Due to their importance in neurons, sodium channels are known molecular targets of neurotoxins and neurotoxicants (Wakeling et al., 2012). There is strong evidence implicating the voltage-sensitive sodium channel as the principal site of insecticidal action of pyrethroids, which has led to extensive studies of the action of pyrethroids on mammalian sodium channels. Binding studies using radioactive pyrethroid demonstrated specific binding of the pyrethroid to rat brain VGSC α subunits (Trainer et al., 1997).</span></span></p>
<p>A prime example of impairments in learning and memory as the adverse outcome for regulatory action is developmental lead exposure and IQ function in children (Bellinger, 2012). Most methods are well established in the published literature and many have been engaged to evaluate the effects of developmental thyroid disruption. The US EPA and OECD Developmental Neurotoxicity (DNT) Guidelines (OCSPP 870.6300 or OECD TG 426) <span style="font-size:16px"><span style="font-family:Arial,Helvetica,sans-serif">as well as OECD TG 443 (OECD, 2018)</span></span> both require testing of learning and memory (USEPA, 1998; OECD, 2007) advising to use the following tests passive avoidance, delayed-matching-to-position for the adult rat and for the infant rat, olfactory conditioning, Morris water maze, Biel or Cincinnati maze, radial arm maze, T-maze, and acquisition and retention of schedule-controlled behaviour. These DNT Guidelines have been deemed valid to identify developmental neurotoxicity and adverse neurodevelopmental outcomes (Makris et al., 2009).</p>
<p>Also, in the frame of the OECD GD 43 (2008) on reproductive toxicity, learning and memory testing may have potential to be applied in the context of developmental neurotoxicity studies. However, many of the learning and memory tasks used in guideline studies may not readily detect subtle impairments in cognitive function associated with modest degrees of developmental thyroid disruption (Gilbert et al., 2012).</p>
adjacentNot SpecifiedNot SpecifiedadjacentNot SpecifiedNot Specified<p><em><strong>Essentiality of MIE and KE1: </strong></em></p>
<p>Evidence from mutation and knockout models demonstrates that perturbation of VGSC function during development impairs nervous system structure and function. Knockout and mutant mouse models of sodium channel α subunits demonstrate varying degrees of adverse outcomes associated with loss or alteration of specific channel subunits. When mRNA for the Nav 1.2 subunit was reduced by approximately 85%, mice exhibited reduced levels of electrical excitability, had high levels of apoptotic neurons in the brainstem and cortex, and died from severe hypoxia within 1–2 days of birth (Planells-Cases et al., 2000). </p>
<p>In insects, only VGSCα are codified. Pyrethroid resistant, or knockdown-resistant houseflies are well known. As this mutation does not alter expression or localisation of the VGSC, it was suspected to alter the affinity of the channel for pyrethroids. Expression of this mutant channel in Xenopus laevis oocytes resulted in VGSCs that were 10-fold less sensitive to cismethirin as assessed using voltage-clamp experiments (Wakeling et al., 2012). </p>
<p>In humans, some mutations have been identified in genes coding for VGSC subunits that result in neuronal hyperexcitability due to subtle changes in channel gating and inactivation (Meisler et al., 2001), these mutations have been linked to various forms of epilepsy (Shafer et al., 2005; Chahine 2018). </p>
<p>Pyrethroids, like these mutations, alter VGSC activation, inactivation and neuronal excitability. However, the mechanisms and magnitude of mutational versus pyrethroid effects are different as well as the duration of the effect. </p>
<p><em><strong>Essentiality of KE1 and KE2: </strong></em></p>
<p>The sodium channel modulator veratridine (VTD) produce the same effect as deltamethrin. In patch recording, this compound rapidly reduced the number of sEPSC without affecting the number of individual burst, but at higher concentration (1 mM) completely removed all sEPSC activity without affecting mEPSC frequency, similar to treatment to TTX (Meyer et al., 2008). Both events – sE(I)PSCs and mE(I)PSCs – are similar in the fact that they occur without any artificial stimulation. The difference between sE(I)PSCs and mE(I)PSCs is coming from the fact that for the sE(I)PSCs there is a chance of action potential-driven events due to intrinsic properties of presynaptic cell and/or network activity. All the mE(I)PSCs, in turn, are recorded in the presence of tetrodotoxin (TTX) which blocks action potential formation and its propagation, therefore mE(I)PSCs are more ‘spontaneous’ events than sE(I)PSCs and can be further used for the quantification of readily releasable pool size. So, it is useful to take both sE(I)PSCs and mE(I)PSCs from the same cell. First, one can record the sE(I)PSCs and then, by adding TTX into the bath solution the mE(I)PSCs. Having both sE(I)PSCs and mE(I)PSCs can help to understand where the changes in synaptic transmission are coming from, i.e. whether it is from the presynaptic side, or postsynaptic or both (Mayer et al., 2008). Titration with tetrodotoxin (TTX) produces a concentration-dependent reduction in the deltamethrin dependent calcium influx, indicating that the alteration in firing rate is consequent to the disruption in the VGSC (Cao et al., 2011).</p>
<p><em><strong>Essentiality of MIE and KE1: </strong></em></p>
<p>Evidence from mutation and knockout models demonstrates that perturbation of VGSC function during development impairs nervous system structure and function. Knockout and mutant mouse models of sodium channel α subunits demonstrate varying degrees of adverse outcomes associated with loss or alteration of specific channel subunits. When mRNA for the Nav 1.2 subunit was reduced by approximately 85%, mice exhibited reduced levels of electrical excitability, had high levels of apoptotic neurons in the brainstem and cortex, and died from severe hypoxia within 1–2 days of birth (Planells-Cases et al., 2000). </p>
<p>In insects, only VGSCα are codified. Pyrethroid resistant, or knockdown-resistant houseflies are well known. As this mutation does not alter expression or localisation of the VGSC, it was suspected to alter the affinity of the channel for pyrethroids. Expression of this mutant channel in Xenopus laevis oocytes resulted in VGSCs that were 10-fold less sensitive to cismethirin as assessed using voltage-clamp experiments (Wakeling et al., 2012). </p>
<p>In humans, some mutations have been identified in genes coding for VGSC subunits that result in neuronal hyperexcitability due to subtle changes in channel gating and inactivation (Meisler et al., 2001), these mutations have been linked to various forms of epilepsy (Shafer et al., 2005; Chahine 2018). </p>
<p>Pyrethroids, like these mutations, alter VGSC activation, inactivation and neuronal excitability. However, the mechanisms and magnitude of mutational versus pyrethroid effects are different as well as the duration of the effect. </p>
<p><em><strong>Essentiality of KE1 and KE2: </strong></em></p>
<p>The sodium channel modulator veratridine (VTD) produce the same effect as deltamethrin. In patch recording, this compound rapidly reduced the number of sEPSC without affecting the number of individual burst, but at higher concentration (1 mM) completely removed all sEPSC activity without affecting mEPSC frequency, similar to treatment to TTX (Meyer et al., 2008). Both events – sE(I)PSCs and mE(I)PSCs – are similar in the fact that they occur without any artificial stimulation. The difference between sE(I)PSCs and mE(I)PSCs is coming from the fact that for the sE(I)PSCs there is a chance of action potential-driven events due to intrinsic properties of presynaptic cell and/or network activity. All the mE(I)PSCs, in turn, are recorded in the presence of tetrodotoxin (TTX) which blocks action potential formation and its propagation, therefore mE(I)PSCs are more ‘spontaneous’ events than sE(I)PSCs and can be further used for the quantification of readily releasable pool size. So, it is useful to take both sE(I)PSCs and mE(I)PSCs from the same cell. First, one can record the sE(I)PSCs and then, by adding TTX into the bath solution the mE(I)PSCs. Having both sE(I)PSCs and mE(I)PSCs can help to understand where the changes in synaptic transmission are coming from, i.e. whether it is from the presynaptic side, or postsynaptic or both (Mayer et al., 2008). Titration with tetrodotoxin (TTX) produces a concentration-dependent reduction in the deltamethrin dependent calcium influx, indicating that the alteration in firing rate is consequent to the disruption in the VGSC (Cao et al., 2011).</p>
High2022-03-15T09:50:562023-11-10T03:34:53