Posttraumatic stress disorder (PTSD) is a severe anxiety disorder characterised by hypothalamic-pituitary-adrenal (HPA)-axis abnormalities, hyperarousal, anxiety, flashbacks of trauma memories and avoidance. Increasing evidence is now accumulating that the disorder is also associated with shrinkage of the hippocampus and cognitive dysfunction that may have its origin in stress-induced excitotoxicity. Animal studies have indeed highlighted a potential role of the excitotoxic glutamatenitric oxide (NO) pathway in the stress response. Since PTSD appears to be an illness that progresses and worsens over time after an initial severe traumatic event, this study has used an animal model that emphasises repeated trauma to investigate the effect of stress on hippocampal NO synthase (NOS) activity, the release of the nitrogen oxide metabolites of NO (NOx), and also the evoked release of cGMP. Furthermore, the modulation and dependency of these responses on glutamate, NO and cGMP activity using drugs selective for these targets, will also be investigated. Rats (n=10/group) were exposed to repeated stress together with saline or drug administration immediately after the stress procedure and continuing for one week post-stress. The animals were then sacrificed for assay of hippocampal NOS activity, NO, and cGMP accumulation. Animals received either the glutamate-NMDA receptor antagonist, memantine (MEM;5mg/kg ip/d), the neuronal NOS selective inhibitor, 7- nitroindazole monosodium salt (7-NINA;20mg/kg ip/d), the cGMP-specific PDE inhibitor, sildenafil (SIL;10mg/kg ip/d) or the NFkb antagonist, pyrollidine dithiocarbamate (PDTC;70mg/kg ip/d). The latter inhibits the nuclear transcription factor, NFkb, responsible for inducing the expression of iNOS, while it also appears to mediate the glutamatergic actions on NOS expression, Stress significantly increased hippocampal NOS activity, as well as significantly increased hippocampal cGMP and NO, levels. These increases were blocked by pretreatment with either PDTC or 7-NINA, while memantine was without effect. Sildenafil significantly augmented stress induced NO, accumulation, as well as cGMP. although the latter failed to reach significance. 7-NINA and memantine significantly blocked the increase in cGMP evoked by time-dependent sensitisation (TDS)-stress, with PDTC attenuating this response, but not significantly. Additionally, administration of each drug separately for seven days without exposure to stress, did not evoke significant changes in NOx levels, compared to the control group. However, significant increases in cGMP levels, compared to the control group, were found with all four drugs. Repeated trauma therefore activates the NO-cGMP pathway, possibly involving actions on both nNOS and iNOS. The NMDA receptor appears less involved after chronic repeated stress, and may have limited therapeutic implications. Sub-cellular NO-modulation, however, may represent an important therapeutic strategy in preventing the effects of severe stress and in treating PTSD. / Thesis (M.Sc. (Pharmacology))--North-West University, Potchefstroom Campus, 2005.

Posttraumatic stress disorder (PTSD) is a severe anxiety disorder characterised by hypothalamic-pituitary-adrenal (HPA)-axis abnormalities, hyperarousal, anxiety, flashbacks of trauma memories and avoidance. Increasing evidence is now accumulating that the disorder is also associated with shrinkage of the hippocampus and cognitive dysfunction that may have its origin in stress-induced excitotoxicity. Animal studies have indeed highlighted a potential role of the excitotoxic glutamatenitric oxide (NO) pathway in the stress response. Since PTSD appears to be an illness that progresses and worsens over time after an initial severe traumatic event, this study has used an animal model that emphasises repeated trauma to investigate the effect of stress on hippocampal NO synthase (NOS) activity, the release of the nitrogen oxide metabolites of NO (NOx), and also the evoked release of cGMP. Furthermore, the modulation and dependency of these responses on glutamate, NO and cGMP activity using drugs selective for these targets, will also be investigated. Rats (n=10/group) were exposed to repeated stress together with saline or drug administration immediately after the stress procedure and continuing for one week post-stress. The animals were then sacrificed for assay of hippocampal NOS activity, NO, and cGMP accumulation. Animals received either the glutamate-NMDA receptor antagonist, memantine (MEM;5mg/kg ip/d), the neuronal NOS selective inhibitor, 7- nitroindazole monosodium salt (7-NINA;20mg/kg ip/d), the cGMP-specific PDE inhibitor, sildenafil (SIL;10mg/kg ip/d) or the NFkb antagonist, pyrollidine dithiocarbamate (PDTC;70mg/kg ip/d). The latter inhibits the nuclear transcription factor, NFkb, responsible for inducing the expression of iNOS, while it also appears to mediate the glutamatergic actions on NOS expression, Stress significantly increased hippocampal NOS activity, as well as significantly increased hippocampal cGMP and NO, levels. These increases were blocked by pretreatment with either PDTC or 7-NINA, while memantine was without effect. Sildenafil significantly augmented stress induced NO, accumulation, as well as cGMP. although the latter failed to reach significance. 7-NINA and memantine significantly blocked the increase in cGMP evoked by time-dependent sensitisation (TDS)-stress, with PDTC attenuating this response, but not significantly. Additionally, administration of each drug separately for seven days without exposure to stress, did not evoke significant changes in NOx levels, compared to the control group. However, significant increases in cGMP levels, compared to the control group, were found with all four drugs. Repeated trauma therefore activates the NO-cGMP pathway, possibly involving actions on both nNOS and iNOS. The NMDA receptor appears less involved after chronic repeated stress, and may have limited therapeutic implications. Sub-cellular NO-modulation, however, may represent an important therapeutic strategy in preventing the effects of severe stress and in treating PTSD. / Thesis (M.Sc. (Pharmacology))--North-West University, Potchefstroom Campus, 2005.

The mammalian spinal cord contains the neuronal circuitry necessary to generate rhythmic locomotor activity in the absence of inputs from the higher brain centre or sensory system. This circuitry is regulated by local neuromodulatory inputs, which can adjust the strength and timing of locomotor output. The free radical gas nitric oxide has been shown to act as an important neuromodulator of spinal circuits, which control locomotion in other vertebrate models such as the tadpole and lamprey. Despite this, the involvement of the NO-mediated soluble guanylate cyclase/cyclic guanosine monophosphate secondary messenger-signalling pathway (NO/sGC/cGMP) in mammalian locomotion has largely been under-investigated. The NADPH diaphorase histochemical reaction was used to identify sources of NO in the lumbar spinal cord. The largest population NADPH diaphorase reactive neurons were located in the dorsal horn, followed by the laminae of the ventral horn, particularly around the central canal (lamina X) and lamina VII. NADPH diaphorase reactive neurons were found along a rostrocaudal gradient between lumbar segments L1 to L5. These results show that that discrete neuronal sources of NO are present in the developing mouse spinal cord, and that these cells increase in number during the developmental period postnatal day P1 – P12. NADPH diaphorase was subsequently used to identify NADPH diaphorase reactive neurons at P12 in the mouse model of ALS using the SODG93A transgenic mouse. Physiological recordings of ventral root output were made to assess the contribution of NO to the regulation induced rhythmic fictive locomotion in the in vitro isolated spinal cord preparation. Exogenous NO inhibits central pattern generator (CPG) output while facilitating and inhibiting motor neuron output at low and high concentrations respectively. Removal of endogenous NO increases CPG output while decreasing motor neuron output and these effects are mediated by cGMP. These data suggest that an endogenous tone of NO is involved in the regulation of fictive locomotion and that this involves the NO/sGC/cGMP pathway. Intracellular recordings from presumed motor neurons and a heterogeneous, unidentified sample of interneurons shows that NO modulates the intrinsic properties of spinal neurons. These data suggest that the net effect of NO appears to be a reduction in motor neuron excitability.