Key Event Title
|Level of Biological Organization|
Key Event Components
Key Event Overview
AOPs Including This Key Event
|AOP Name||Role of event in AOP|
|Binding of antagonist to NMDARs impairs cognition||KeyEvent|
|nAChR activation - colony death/failure2||KeyEvent|
|nAChR activation - colony loss 8||KeyEvent|
|NIS inhibition and learning and memory impairment||KeyEvent|
|Oxidative stress and Developmental Neurotoxicity||KeyEvent|
|During brain development||High|
Key Event Description
Biological state: There are striking differences in neuronal network formation and function among the developing and mature brain. The developing brain shows a slow maturation and a transient passage from spontaneous, long-duration action potentials to synaptically-triggered, short-duration action potentials.
Furthermore, at this precise developmental stage the neuronal network is characterised by "hyperexcitability”, which is related to the increased number of local circuit recurrent excitatory synapses and the lack of γ-amino-butyric acid A (GABAA)-mediated inhibitory function that appears much later. This “hyperexcitability” disappears with maturation when pairing of the pre- and postsynaptic partners occurs and synapses are formed generating population of postsynaptic potentials and population of spikes followed by developmental GABA switch. Glutamatergic neurotransmission is dominant at early stages of development and NMDA receptor-mediated synaptic currents are far more times longer than those in maturation, allowing more calcium to enter the neurons. The processes that are involved in increased calcium influx and the subsequent intracellular events seem to play a critical role in establishment of wiring of neural circuits and strengthening of synaptic connections during development (reviewed in Erecinska et al., 2004). Neurons that do not receive glutaminergic stimulation are undergoing developmental apoptosis.
During the neonatal period, the brain is subject to profound alterations in neuronal circuitry due to high levels of synaptogenesis and gliogenesis. For example, in neuroendocrine regions such as the preoptic area-anterior hypothalamus (POA-AH), the site of gonadotropin-releasing hormone (GnRH) system is developmentally regulated by glutamatergic neurons. The changes in the expression of the N-methyl-D-aspartate (NMDA) receptor subunits NR1 and NR2B system begin early in postnatal development, before the onset of puberty, thereby playing a role in establishing the appropriate environment for the subsequent maturation of GnRH neurons (Adams et al., 1999).
Biological compartments: Neural network formation and function happen in all brain regions but it appears to onset at different time points of development (reviewed in Erecinska et al., 2004). Glutamatergic neurotransmission in hippocampus is poorly developed at birth. Initially, NMDA receptors play important role but the vast majority of these premature glutamatergic synapses are “silent” possibly due to delayed development of hippocampal AMPA receptors. In contrast, in the cerebral cortex the maturation of excitatory glutamatergic neurotransmission happens much earlier. The “silent” synapses disappear by PND 7-8 in both brain regions mentioned above.
There is strong evidence suggesting that NMDA receptor subunit composition controls synaptogenesis and synapse stabilization (Gambrill and Barria, 2011). It is established fact that during early postnatal development in the rat hippocampus, synaptogenesis occurs in parallel with a developmental switch in the subunit composition of NMDA receptors from NR2B to NR2A. It is suggested that early expression of NR2A in organotypic hippocampal slices reduces the number of synapses and the volume and dynamics of spines. In contrast, overexpression of NR2B does not affect the normal number and growth of synapses. However, it does increase spine motility, adding and retracting spines at a higher rate. The C terminus of NR2B, and specifically its ability to bind CaMKII, is sufficient to allow proper synapse formation and maturation. Conversely, the C terminus of NR2A was sufficient to stop the development of synapse number and spine growth. These results indicate that the ratio of synaptic NR2B over NR2A controls spine motility and synaptogenesis, and suggest a structural role for the intracellular C terminus of NR2 in recruiting the signalling and scaffolding molecules necessary for proper synaptogenesis. Interestingly, it was found that genetic deletion of NR3A accelerates glutamatergic synaptic transmission, as measured by AMPAR-mediated postsynaptic currents recorded in hippocampal CA1. Consistent, the deletion of NR3A accelerates the expression of the glutamate receptor subunits NR1, NR2A, and GluR1 sugesting that glutamatergic synapse maturation is critically dependent upon activation of NMDA-type glutamate receptors (Henson et al., 2012).
General role in biology: The development of neuronal networks can be distinguished into two phases: an early ‘establishment’ phase of neuronal connections, where activity-dependent and independent mechanisms could operate, and a later ‘maintenance’ phase, which appears to be controlled by neuronal activity (Yuste and Sur, 1999). These neuronal networks facilitate information flow that is necessary to produce complex behaviors, including learning and memory (Mayford et al., 2012).
How It Is Measured or Detected
Methods that have been previously reviewed and approved by a recognized authority should be included in the Overview section above. All other methods, including those well established in the published literature, should be described here. Consider the following criteria when describing each method: 1. Is the assay fit for purpose? 2. Is the assay directly or indirectly (i.e. a surrogate) related to a key event relevant to the final adverse effect in question? 3. Is the assay repeatable? 4. Is the assay reproducible?
In vivo: The recording of brain activity by using electroencephalography (EEG), electrocorticography (ECoG) and local field potentials (LFP) assists towards the collection of signals generated by multiple neuronal cell networks. Advances in computer technology have allowed quantification of the EEG and expansion of quantitative EEG (qEEG) analysis providing a sensitive tool for time-course studies of different compounds acting on neuronal networks' function (Binienda et al., 2011). The number of excitatory or inhibitory synapses can be functionally studied at an electrophysiological level by examining the contribution of glutamatergic and GABAergic synaptic inputs. The number of them can be determined by variably clamping the membrane potential and recording excitatory and inhibitory postsynaptic currents (EPSCs or IPSCs) (Liu, 2004).
In vitro: 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). Patch clamping technique can also be used to measure neuronal network activity.
Domain of Applicability
In vitro studies in brain slices applying electrophysiological techniques showed significant variability among species (immature rats, rabbits and kittens) related to synaptic latency, duration, amplitude and efficacy in spike initiation (reviewed in Erecinska et al., 2004).
Adams MM, Flagg RA, Gore AC., Perinatal changes in hypothalamic N-methyl-D-aspartate receptors and their relationship to gonadotropin-releasing hormone neurons. Endocrinology. 1999 May;140(5):2288-96.
Binienda ZK, Beaudoin MA, Thorn BT, Ali SF. (2011) Analysis of electrical brain waves in neurotoxicology: γ-hydroxybutyrate. Curr Neuropharmacol. 9: 236-239.
Erecinska M, Cherian S, Silver IA. (2004) Energy metabolism in mammalian brain during development. Prog Neurobiol. 73: 397-445.
Gambrill AC, Barria A. NMDA receptor subunit composition controls synaptogenesis and synapse stabilization. Proc Natl Acad Sci U S A. 2011:108(14):5855-60.
Gopal K. (2003) Neurotoxic effects of mercury on auditory cortex networks growing on microelectrode arrays: a preliminary analysis. Neurotoxicol Teratol. 25: 69-76.
Gramowski A, Schiffmann D, Gross GW. (2000) Quantification of acute neurotoxic effects of trimethyltin using neuronal networks cultures on microelectrode arrays. Neurotoxicology 21: 331-342.
Henson MA, Larsen RS, Lawson SN, Pérez-Otaño I, Nakanishi N, Lipton SA, Philpot BD. (2012) Genetic deletion of NR3A accelerates glutamatergic synapse maturation. PLoS One. 7(8).
Hogberg HT, Sobanski T, Novellino A, Whelan M, Weiss DG, Bal-Price AK. (2011) Application of micro-electrode arrays (MEAs) as an emerging technology for developmental neurotoxicity: evaluation of domoic acid-induced effects in primary cultures of rat cortical neurons. Neurotoxicology 32: 158-168.
Johnstone AFM, Gross GW, Weiss D, Schroeder O, Shafer TJ. (2010) Use of microelectrode arrays for neurotoxicity testing in the 21st century Neurotoxicology 31: 331-350.
Keefer E, Norton S, Boyle N, Talesa V, Gross G. (2001) Acute toxicity screening of novel AChE inhibitors using neuronal networks on microelectrode arrays. Neurotoxicology 22: 3-12.
Liu G. (2004) Local structural balance and functional interaction of excitatory and inhibitory synapses in hippocampal dendrites. Nat Neurosci. 7: 373-379.
Mayford M, Siegelbaum SA, Kandel ER. (2012) Synapses and memory storage. Cold Spring Harb Perspect Biol. 4. pii: a005751.
McConnell ER, McClain MA, Ross J, LeFew WR, Shafer TJ. (2012) Evaluation of multi-well microelectrode arrays for neurotoxicity screening using a chemical training set. Neurotoxicology 33: 1048-1057.
Novellino A, Scelfo B, Palosaari T, Price A, Sobanski T, Shafer TJ, Johnstone AF, Gross GW, Gramowski A, Schroeder O, Jügelt K, Chiappalone M, Benfenati F, Martinoia S, Tedesco MT, Defranchi E, D'Angelo P, Whelan M. (2011) Development of micro-electrode array based tests for neurotoxicity: assessment of interlaboratory reproducibility with neuroactive chemicals. Front Neuroeng. 4: 4.
Yuste R, Peinado A, Katz LC. (1992) Neuronal domains in developing neocortex. Science 257: 665-669.