While there is clear evidence for sensory neuron expression of GPR54, receptor for kisspeptin, and possible role of kisspeptin in pain modulation (1), there is no study on the effect and mechanism(s) of kisspeptin on sensory neurons. hence, the aim of this study was to investigate the effects and mechanism of kisspeptin on intracellular free calcium levels in cultured rat dorsal root ganglion (DRG) neurons. The 2 day-old Sprague-Dawley rat pups were killed by decapitation and primary cultures of dissociated DRG neurons from all levels of spinal cord were prepared by enzymatic and mechanical methods. DRG neurons were loaded with 1 uM Fura-2 AM and intracellular free calcium responses were quantified by the changes in 340/380 ratio by digital imaging microscopy, with excitation at 340 and 380 nm and emission at 500 nm. Data (normalised and expressed as mean ± SEM) were analysed using the unpaired Student’s t-test, P <.05 defining statistical significance. Bath application of kisspeptin at concentrations of 100 nM and 1 uM increased intracellular free calcium levels from basal values of 100±0.0% to 115.9±3.5% (n=24, p<0.001) and 100±0.0% to 130.9± 3.1% (n=42, p<0.001), respectively. In the absence of extracellular calcium, the kisspeptin (1 uM) induced attenuated but still significant increase in intracellular free calcium levels; basal values of 100±0.0% to 108.1±3.6 (n=23, p<0.01). The kisspeptin-induced [intracellular free calcium responses persisted in the presence of PKC inhibitor chelerythrine chloride (10 uM, n=27). Data from this study revealed for the first time that kisspeptin activates intracellular calcium signalling in cultured rat DRG neurons in a dose dependent manner, and this activation is independent of protein kinase C.