Endocrine Abstracts

Orexins, the novel pluripotent hypothalamic peptides, have been shown to exert important roles in the regulation of multiple physiological functions and behaviors including appetite, sleep and wakefulness and energy homeostasis through neural and endocrine mechanisms. Evidence emerging from recent research indicates that orexins may be involved in many other physiological functions. Our previous results have suggested that orexin-A and B activate Ca²⁺ signaling in cultured rat dorsal root ganglion (DRG) neurons implicating a role in nociception, and the aim of the present study was to investigate whether this orexin receptors mediated signaling involves PKC pathways in this sensory neurones. Following enzymatic digestion and mechanical agitation the DRG neurons were cultured on coated coverslips and loaded with 1 μmol Fura-2 AM. [Ca²⁺]_i responses were quantified by the changes in 340/380 ratio for individual DRG neurons using the imaging system consisting of CCD camera coupled to an inverted microscope with a 40× (1.30 NA) objective. All data were analyzed by using unpaired t test, P<0.05 defining statistical significance. The non-peptide OX₁ selective receptor antagonist SB-334867-A (1 μM) inhibited the orexin-A (200 nM) and orexin-B (200 nM)-induced calcium responses (57.2±4.2% versus orexin-A, n=5, and 65.9±3.6% versus orexin-B, n=9). The PKC inhibitor chelerythrine chloride also decreased the orexin-A (200 nM)-induced calcium responses (59±5.2% and 5±1.7% versus orexin-A, n=7 for both 10 and 100 μM). In conclusion, the results suggest that ORX A and -B cause an increase in free intracellular calcium through PKC pathway activation, which could be associated with nociceptive modulation and pain.