Effect Of Progesterone On The Change Of Free Ca²⁺ Concentration In Sensory Neurons Of Newborn Rabbit Induced By ATP In Vitro

Progesterone(progesterone, PROG) was required to maintain the pregnancy as an important steroid hormones. With further study found that the nervous system can synthesize and secrete progesterone, which affect the function of the nervous system, was an important signaling molecule of the nervous system. Clinically, there is growing concern about treatment of neurological disorderswith estrogen and progesterone. Dorsal root ganglion(DRG) sensory was a first-class in signal, peripheral sensory information collected for preliminary processing, then passed to the hub. Our studies found that the DRG vascular permeability greater than cerebral blood vessels permeability for rabbits, indicating there present an acceptance signaling molecules DRG; in addition, there exist endocrine- immune signaling molecule receptors in the internal of DRG, which means DRG was an important node in the system of neuroendocrine. So DRG has significance for transmission and modulation of a variety of sensory information. Fan Juan et al., found that the progesterone and ATP had impact of changes in DRG neuronal membrane potential. But it is unclear about how the combination of Progesterone and ATP impact neurons intracellular calcium ion of DRG. P2X3 was a member of ATP receptors, DRG mainly express for P2X3 receptor and P2X3 receptor had significantly permeability for Ca2+. In this study, based on the characteristics of distribution, we used double immunofluorescence labeling progesterone receptor(PR) and P2X3 receptor; using MTT method detected of different concentrations of progesterone in vitro activity of DRG neurons; using the confocal laser scanning microscopy techniques monitoring of different concentrations of ATP and progesterone impact on the intracellular calcium ion concentration([Ca2+]i) in DRG neurons at real-time dynamic. The results as follows:(1) When cultured in the third day, the cells were mainly small and medium sized neurons(97.179%), both of which were progesterone receptor(PR) and P2X3 receptor fluorescent double labeled cells.(2) Different concentrations of ATP can caused DRG neurons [Ca2+]i rapid increase quickly, and there had concentration-dependent effect, of which 1000 mmol/L of ATP can be induced in sensory neurons [Ca2+]i standardized fluorescence value(F/F0) reached 1.672, by using SPSS 22.0 single factor variance analysis test had extremely significant difference; 1000 mol/L of ATP stimulation, without calcium and with calcium in cultured solution, sensory neurons in Ca2+ standardized fluorescence value(F/F0) was 1.672(P < 0.01) and 1.304(P < 0.01), and between the two had extremely significant difference(P < 0.01).(3) Progesterone induced calcium ion concentration changes of DRG neuron showed no significant difference, but showed three trends: increase in the concentration of calcium ions, calcium ion concentration did not change significantly, reduce the concentration of calcium ion, which with reduce the calcium ion concentration classes was the main type of neurons, with the increase of the concentration of progesterone by percentage decrease(90.91% and 84.61%, 72.73%, 70.00%).(4) Without calcium and with calcium in cultured solution, with 1000 mmol/L ATP as the stimulation, progesterone had significant effects on the changes of [Ca2+]i in neurons and there had concentration-dependent effect; without calcium and with Ca2+ in culture solution, the same concentration of progesterone group changes on intracellular [Ca2+]i has significant difference.The study showed that the small and medium-sized neurons of DRG exists the effects of progesterone and ATP conditions. ATP increased the intracellular [Ca2+]i mode of the neurons of DRG in the newborn rabbits, which may be supplemented by calcium influx and calcium release. The sensitivity of the small medium-sized neurons of DRG in the rabbit to the progesterone was different. Progesterone does not affect the release of Ca2+ in neurons, progesterone may through progesterone receptor affect the signal transduction pathway of P2X3 receptor and then affect the intracellular calcium release to increase intracellular [Ca2+]i.