Synergistic effect of orexin-glutamate co-administration on spontaneous discharge rate of locus coeruleus neurons in morphine-dependent rats.

Various intrinsic and extrinsic factors can increase the spontaneous discharge rate of locus coeruleus (LC) neurons. Among the extrinsic ones, orexinergic neurons of the lateral hypothalamus (LH) send widespread projections to LC region. Accumulating evidence support the involvement of glutamate in mediating the excitatory effect of orexin-A on LC neurons. In addition, both orexinergic and glutamatergic systems have been shown to play critical roles in molecular changes underlying the development of morphine dependence. The present study was designed to investigate the interaction between orexin-A and glutamate in modulating the firing rate of LC neurons. Regarding the role of both orexinergic and glutamatergic systems in morphine dependence, this effect was also investigated in morphine dependent rats. For this purpose, spontaneous discharge rate of LC neurons was recorded using the whole-cell patch clamp recording method in presence of orexin-A, glutamate or orexin-A plus glutamate in acutely prepared brain slices. Results indicated that superfusion of either orexin-A or glutamate enhances the firing rate of LC neurons in both dependent and non-dependent rats. However, co-application of orexin-A and glutamate elicited a significant synergism in enhancement of firing rate only in morphine dependent animals. In conclusion, it seems that development of morphine dependence promotes adaptations in locus coeruleus neurons that potentiate the orexin-glutamate interaction.

Intra-LC microinjection of orexin type-1 receptor antagonist SB-334867 attenuates the expression of glutamate-induced opiate withdrawal like signs during the active phase in rats.

Opiate withdrawal syndrome is temporally associated with the hyperactivity of locus coeruleus neurons. Previous studies have shown that this hyperactivity, at least in part, results from the activity of excitatory afferents which mainly include the orexinergic neurons of hypothalamus and glutamatergic neurons of paragigantocellularis (PGi) nucleus. The effect of intra LC orexin type 1 receptor antagonism was investigated on expression of glutamate-induced morphine withdrawal-like signs in rats. Regarding the involvement of both orexin and LC in modulation of circadian rhythm, experimental procedures were performed during the rest (day) and the active (night) phases. Male Wistar rats (250-300g) received escalating doses (6, 16, 26, 36, 46, 56, 66mg/kg, 2ml/kg) of morphine sulfate subcutaneously for 7days. Then, glutamate (100nM, 200nl) was microinjected into the LC region and the subsequent behavioral manifestations were visually monitored in both rest and active phases. SB-334867 (as a selective orexin type 1 receptor antagonist) was microinjected into the LC prior to glutamate administration. Results indicate that intra-LC microinjection of glutamate elicits morphine withdrawal-like behavioral signs in rats. It is noteworthy that this effect was significantly suppressed in rats pretreated with SB-334867 only during the active phase. It could be concluded that orexin-A plays a role in expression of glutamate-induced opiate withdrawal-like signs and differential orexinergic tone during the rest and active phases might explain the observed difference in activity of LC neurons.

The role of endogenous hydrogen sulfide in pathogenesis of chronotropic dysfunction in rats with cirrhosis.

Endogenous hydrogen sulfide is produced by cystathionine-γ-lyase and cystathionine-ß-synthase in a variety of tissues and has recently been implicated in the regulation of cardiac functions. Acceleration of the heart rate in response to catecholamines is impaired in patients with cirrhosis. The present study was aimed to examine the role of endogenous hydrogen sulfide in the pathogenesis of chronotropic dysfunction in rats with cirrhosis. Cirrhosis was induced by surgical ligation of bile duct in rats. There was no significant difference in atrial cystathionine-γ-lyase and cystathionine-ß-synthase mRNA levels in control and cirrhotic rats as assessed by quantitative RT-PCR. Four weeks after bile duct ligation or sham surgery the atria were isolated and chronotropic responsiveness to adrenergic stimulation was assessed using standard organ bath. Incubation of the atria with propargylglycine (PAG, a cystathionine-γ-lyase inhibitor) and amino-oxyacetic acid (AOAA, a cystathionine-ß-synthase inhibitor) was associated with a significant desensitization of chronotropic response to adrenergic stimulation in controls rats. This indicates that endogenous hydrogen sulfide might be involved in modulation of adrenergic signaling in the atrium. Bile duct ligation was associated with impaired chronotropic responsiveness to adrenergic stimulation in comparison with sham-operated rats. In contrast to control group, incubation of the atria with PAG and AOAA was able to partially improve the chronotropic responsiveness to adrenergic stimulation in cirrhotic rats. Our data shows that local inhibition of endogenous hydrogen sulfide in atria has opposite effect in cirrhotic versus control rats and may play a role in physiological modulation of adrenergic signaling in the atrium.