Nitric Oxide Signaling Strengthens Inhibitory Synapses of Cerebellar Molecular Layer Interneurons through a GABARAP-Dependent Mechanism.

Nitric oxide (NO) is an important signaling molecule that fulfills diverse functional roles as a neurotransmitter or diffusible second messenger in the developing and adult CNS. Although the impact of NO on different behaviors such as movement, sleep, learning, and memory has been well documented, the identity of its molecular and cellular targets is still an area of ongoing investigation. Here, we identify a novel role for NO in strengthening inhibitory GABAA receptor-mediated transmission in molecular layer interneurons of the mouse cerebellum. NO levels are elevated by the activity of neuronal NO synthase (nNOS) following Ca2+ entry through extrasynaptic NMDA-type ionotropic glutamate receptors (NMDARs). NO activates protein kinase G with the subsequent production of cGMP, which prompts the stimulation of NADPH oxidase and protein kinase C (PKC). The activation of PKC promotes the selective strengthening of α3-containing GABAARs synapses through a GΑΒΑ receptor-associated protein-dependent mechanism. Given the widespread but cell type-specific expression of the NMDAR/nNOS complex in the mammalian brain, our data suggest that NMDARs may uniquely strengthen inhibitory GABAergic transmission in these cells through a novel NO-mediated pathway.SIGNIFICANCE STATEMENT Long-term changes in the efficacy of GABAergic transmission is mediated by multiple presynaptic and postsynaptic mechanisms. A prominent pathway involves crosstalk between excitatory and inhibitory synapses whereby Ca2+-entering through postsynaptic NMDARs promotes the recruitment and strengthening of GABAA receptor synapses via Ca2+/calmodulin-dependent protein kinase II. Although Ca2+ transport by NMDARs is also tightly coupled to nNOS activity and NO production, it has yet to be determined whether this pathway affects inhibitory synapses. Here, we show that activation of NMDARs trigger a NO-dependent pathway that strengthens inhibitory GABAergic synapses of cerebellar molecular layer interneurons. Given the widespread expression of NMDARs and nNOS in the mammalian brain, we speculate that NO control of GABAergic synapse efficacy may be more widespread than has been appreciated.

Drug-Induced Seizures: Considerations for Underlying Molecular Mechanisms.

A broad spectrum of chemical entities have been associated with drug-induced seizure (DIS), emphasizing the importance of this potential liability across various drug classes (e.g., antidepressants, antipsychotics, antibiotics, and analgesics among others). Despite its importance within drug safety testing, an understanding of the molecular mechanisms associated with DIS is often lacking. The etiology of DIS is understood to be a result of either a deficit in inhibitory (e.g., gamma aminobutyric acid) or an elevated excitatory (e.g., glutamate) signaling, leading to synchronous neuronal depolarization affecting various brain regions and impairing normal neurological functions. How this altered neuronal signaling occurs and how these changes interact with other non-brain receptor driven DIS-associated changes such as metabolic disturbances, electrolyte imbalances, altered drug metabolism, and withdrawal effects are poorly understood. Herein, we discuss important molecular mechanisms identified in DIS for several drugs and/or drug classes. With a better understanding of the molecular mechanisms associated with DIS, in vivo or in vitro models may be applied to characterize and mitigate DIS risk during drug development. Susceptibility stratification for DIS presents species differences in the following order beagle dogs > rodents and cynomolgus monkeys > Göttingen minipigs with a more than 2-fold difference between canines and minipigs, which is important to consider during non-clinical species selection. While clinical signs such as myoclonus, severe muscle jerks, or convulsions are often associated with abnormal epileptiform EEG activity, tremors are most of the time physiological and rarely observed with concurrent epileptiform EEG activity which need to be considered during DIS risk evaluation.