The hypothalamic peptides orexin A/hypocretin-1 and Orexin B/hypocretin-2 are involved in a range of physiological functions including control of feeding and energy metabolism, sleep and arousal. Orexin fibers innervate many regions of the brain and spinal cord which include areas involved in pain processing and emerging evidence suggests that orexins modulate pain transmission. This study investigate the effects of Orexin A on intracellular calcium, ([Ca2+]i), signals in cultured rat dorsal root ganglion (DRG) neurons, with the aim of exploring possible involvement of this agent in nociceptive transmission. DRG neuronal cultures were loaded with 1 μmol Fura-2 AM and Ca2+ responses were assessed by using the fluorescent ratiometry. Fura-2 loaded DRG cultures were excited at 340 and 380 nm, and emission was recorded at 510 nm by using imaging system consisting of CCD camera coupled to an inverted microscope with a 40x (1.30 NA S Fluor, Oil) objective. [Ca2+]i changes were determined by the change in 340/380 ratio (basal-peak) was also calculated for individual DRG neurons in selected microscopic fields. All data were analyzed by using unpaired t test, with a 2-tailed P level of <.05 defining statistical significance. ORX-A caused increase in [Ca2+]i in a dose dependent manner. The ORX-A-induced [Ca2+]i responses were similar to those observed with high-K+ (30 mM). The mean 340/380 nm ratios were (baseline vs OrX-A): 0.90±0.01 vs 1.34±0.03 (1 nM OrX-A, P<0.001, n=21); 1.10±0.03 vs. 2.15±0.09 (10 nM OrX-A, P<0.001, n=21); 0.82±0.02 vs. 1.65±0.09 (100 nM OrX-A, P<0.001, n=18) and 0.72±0.02 vs. 1.69±0.08 (200 nM OrX-A, P<0.001, n=34), respectively. These results, in line with previous findings in different preparations, show that Orexin A increases intracellular calcium levels in a dose-dependent fashion in DRG neurons. We conclude that orexin-A has excitatory effects on DRG neurones, consistent with the perspective that orexin/hypocretins have a role in orchestrating reactions related to nociception, pain and temperature sense. Keywords: Orexin A; fluorescence calcium imaging, nociception, sensory neurones