Regulation Of Limbic System-mediated Memory And Behavior By Gaseous Signal Molecule

Part Ⅰ Effects and mechanisms of gas molecule nitric oxide on addiction-related memory in rats during cocaine withdrawalAim:As an endogenous gas molecule, nitric oxide (NO) exerts a host of biological effects in organisms. Numerous studies have shown a role for NO in cocaine addiction. However, little is known about the effect of NO in neurobehavioral adaptations after cocaine withdrawal. Previous studies have reported that the function of accumbal α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor is upregulated by cocaine withdrawal, which is believed to contribute to behavioral adaptations to cocaine. Considering that NO can regulate surface expression of AMPA receptors by the manner of S-nitrosylation, we postulated that NO might participate in the formation of neurobehavioral adaptations during cocaine withdrawal.Methods:Protein cross-linking assay was adopted to investigate surface expressions of GluR1and GluR2subunits. Whole-cell patch-clamp and field potentials recording were used to examine the AMPA receptor-mediated EPSC (AMPAR-EPSC) and long-term depression (LTD). Behavioral experiments including locomotor activity and sucrose preference were adopted to investigate animals’ reward response.Results:We showed that cocaine withdrawal led to disproportionate upregulation of surface GluRl and GluR2subunits. Further increase in surface GluR2by blocking endocytosis or promoting surface insertion restored this synaptic adaptation and prevented the expression of locomotor sensitization. Moreover, the nitric oxide (NO) content and S-nitrosylated N-ethylmaleimide sensitive factor (NSF) level were increased during withdrawal. On the other side, exogenous NO promoted surface GluR2insertion via S-nitrosylation of NSF and blocked the expression of behavioral sensitization.Conclusion:These results suggest that the upregulation of surface GluR2is a compensatory mechanism to limit long-lasting maladaptive behavioral responses to cocaine, and S-nitrosylated accumbal NSF could reverse neurobehavioral adaptations to cocaine. Part Ⅱ Effects and mechanisms of endogenous hydrogen sulfide on hippocampal learning and memory in ratsAim:Hydrogen sulfide (H2S), an endogenous gas molecule, exerts a plenty of biological effects in organisms. Given that H2S in physiological concentrations can facilitate NMDA receptor (NMDAR)-dependent long-term potentiation (LTP), we hypothesized that H2S elevations might improve, whereas reductions might aggravate, cognitive functions.Methods:H2S content in hippocampus tissue sample was measured using methylene blue method. Contextual fear conditioning and novel object recognition were adopted to test hippocampus-dependent cognition. Electrophysiological methods (whole cell patch-clamp and field potentials recording) were used to examine AMPAR-EPSC and LTP. Western blotting was also used to investigate protein expressions in this section. Results:We found that H2S level was significantly elevated in the hippocampus of rats following fear conditioning. Inhibition of cystathionine-β-synthase (CBS), the predominant H2S-producing enzyme in the brain, prevented training-induced hippocampal H2S elevation and impaired contextual fear memory. Whereas exogenous administration of H2S donor dose-dependently enhanced hippocampus-dependent contextual fear memory. Similarly, rats treated with H2S administration displayed an enhanced ability in novel object recognition and treatment with CBS inhibitor impaired this recognition. Further experiments show that H2S selectively enhanced NR2A-containing NMDA receptor currents and LTP, and blockade of NR2A-containing NMDA receptor eliminated the pro-cognitive effects of H2S in vitro and in vivo. Moreover, H2S-mediated effects may also correlate with the activation of signaling pathways such as PKA, PKC, CaMKII and CREB.Conclusion:These studies show that a pivotal role for H2S in hippocampal memory, providing information for further understanding the physiological functions of H2S in the center nervous system. Part III Effects and mechanisms of gas molecule hydrogen sulfide on amygdala-mediated emotional memory in ratsAim:Amygdala is a key brain region that performs a primary role in the processing and memory of emotional reactions. Previous studies have shown dysfunctions of amygdala-mediated emotional memory in frontotemporal lobar degeneration and Alzheimer’s disease, and emotional disorders in autism. As the third gaseous messenger molecule, hydrogen sulfide (H2S) serves as an endogenous neuromodulator in the hippocampus. However, little is known about whether H2S has a role in amygdala-mediated emotional memory and synaptic plasticity.Methods:H2S content in brain tissue sample was measured using methylene blue method. Cued fear conditioning and conditioned taste aversion (CTA) were adopted to test emotional memory. Electrophysiological methods (whole cell patch-clamp and field potentials recording) were used to examine AMPAR-EPSC and LTP in the amygdale. Protein cross-linking assay and western blotting were used to investigate the total and surface protein expressions in this section.Results:It was found that H2S level was significantly elevated in the brain of rats following fear conditioning. Exogenous administration of H2S donor dose-dependently enhanced amygdala-dependent cued fear memory and prevented memory forgetting. Furthermore, H2S treatment attenuated extinction learning and the retention of extinction memory, promoted spontaneous recovery of fear memory and reinstatement. Similarly, exogenous H2S treatment significantly improved the memory ability of conditioned taste aversion, another type of amygdala-mediated emotional memory. Immunochemical staining showed extensive expression of cystathionine-β-synthase (CBS), the predominant H2S-producing enzyme, in the amygdala. Infusion with CBS inhibitor into lateral amygdala before fear training impaired cued memory. Moreover, H2S treatment significantly increased dendrite spine density, CREB activity, BDNF content and surface NR2B expression in the lateral amygdala. Bath application of H2S donor selectively increased NR2B-mediated current in the thalamus-amygdala pathway and the enhancement of NMDA receptor-dependent LTP by H2S required the activation of NR2B-containing NMDA receptor. Moreover, lateral amygdala infusion of NR2B-selective NMDA receptor antagonist eliminated the memory enhancement of H2S during cued test.Conclusion:These studies show that H2S has ability to promote emotional memory via regulating the function of NR2B-containing NMDA receptor, deepening the understanding of physiological functions of H2S in the center nervous system and suggesting that H2S-releasing agents might be a new strategy for the treatment of emotional disorders.