Expression of Glial Cell Line-Derived Neurotrophic Factor (GDNF) and the GDNF Family Receptor Alpha Subunit 1 in the Paravaginal Ganglia of Nulliparous and Primiparous Rabbits / 대한배뇨장애요실금학회지

PURPOSE: To evaluate the expression of glial cell line-derived neurotrophic factor (GDNF) and its receptor, GDNF family receptor alpha subunit 1 (GFRα-1) in the pelvic (middle third) vagina and, particularly, in the paravaginal ganglia of nulliparous and primiparous rabbits. METHODS: Chinchilla-breed female rabbits were used. Primiparas were killed on postpartum day 3 and nulliparas upon reaching a similar age. The vaginal tracts were processed for histological analyses or frozen for Western blot assays. We measured the ganglionic area, the Abercrombie-corrected number of paravaginal neurons, the cross-sectional area of the neuronal somata, and the number of satellite glial cells (SGCs) per neuron. The relative expression of both GDNF and GFRα-1 were assessed by Western blotting, and the immunostaining was semiquantitated. Unpaired two-tailed Student t -test or Wilcoxon test was used to identify statistically significant differences (P≤0.05) between the groups. RESULTS: Our findings demonstrated that the ganglionic area, neuronal soma size, Abercrombie-corrected number of neurons, and number of SGCs per neuron were similar in nulliparas and primiparas. The relative expression of both GDNF and GFRα-1 was similar. Immunostaining for both GDNF and GFRα-1 was observed in several vaginal layers, and no differences were detected regarding GDNF and GFRα-1 immunostaining between the 2 groups. In the paravaginal ganglia, the expression of GDNF was increased in neurons, while that of GFRα-1 was augmented in the SGCs of primiparous rabbits. CONCLUSIONS: The present findings suggest an ongoing regenerative process related to the recovery of neuronal soma size in the paravaginal ganglia, in which GDNF and GFRα-1 could be involved in cross-talk between neurons and SGCs.

Feasibility study of transplantation of penile corpus cavernosum and major pelvic ganglion in renal subserous region / 北京大学学报(医学版)

Objective:To study the feasibility of transplantation of normal rat penile corpus cavernosum and major pelvic ganglion (MPG)into the renal subserous region of a Nu /Nu mouse based on allograft technology.Methods:Penile corpus cavernosum and MPG,harvested from Sprague-Dawley (SD)rats under sterile condition,were transplanted underneath the kidney capsule of Nu /Nu mice through the mi-crosurgery instruments and surgery microscope.The histopathologic changes and cellular proliferation in the transplanted penile corpus cavernosum and MPG were then analyzed at the end of 1week and 4 weeks after transplantation.Histological staining and immunohistochemical staining were used to evaluate the main outcome measures.Results:After 1 week,the tissue morphology of the transplanted corpus caverno-sum underneath the kidney capsule of Nu /Nu mice was consistent with normal penile corpus cavernosum, and blood could be observed in the penis cavernous sinus of the graft;after 4 weeks,the mophorlogy of the tranplanted corpus cavernosum near the kidney was consistent with normal penile corpus cavernosum, while fibrosis was noteworthy in the graft away from the kidney,but blood could still be seen in the penis cavernous sinus.After 1 week,the tissue morphology of the transplanted MPG was consistent with normal MPG,multiple islet-like cell clusters could be seen in the transplanted MPG in the renal subserous re-gion,and angiogenesis could be observed near the kidney;after 4 weeks,a network of blood vessels was clearly visible away from the kidney,and islet-like cell clusters were still clearly observed in the trans-planted MPG.In addition,ki67 positive cells were observed in the transplanted penile corpus cavernosum and MPG after 4 weeks of transplantation,which indicated that there was still cell proliferation activity in the grafts.Conclusion:The transplanted corpus cavernosum and MPG underneath the kidney capsule of Nu /Nu mice could survive at least 4 weeks.Moreover,the inner structure of the transplanted corpus ca-vernosum and MPG was close to the normal tissue.The underlining mechanism may be related to the lo-cal microenvironment underneath the kidney capsule of Nu /Nu mice and the neovascularization in the transplanted grafts.

Neuronal Excitatory Action of GABA on the Pelvic Ganglia

OBJECTIVE: In the central nervous system, gamma-aminobutyric acid (GABA) is well known to act as an inhibitory neurotransmitter by hyperpolarizing postsynaptic neurons through gating GABA-activated Cl- channels. To date, however, the functional roles of GABA remain unclear in the autonomic nervous system. In the present study, we characterize GABA-activated Cl- currents in the neurons of major pelvic ganglia (MPG). METHODS: MPG neurons, located on the lateral surfaces of the prostate gland, from male rats were enzymatically dissociated. Ionic currents were recorded using whole-cell variant patch-clamp technique. Membrane potential was recorded under current clamp mode. Current traces were filterd at 2kHz by using 4-pole Bassel filter in the amplifier. RESULTS: Application of GABA (100micrometer) induced inward currents in the neurons, with holding potentials being maintained below the Cl- equilibrium potential (ECl). The GABA response was concentration-dependent and its reversal potential was close to the theoretical ECl. The GABA-induced Cl- currents were largely blocked by bicuculline (10micrometer, n=5), a GABAA receptor antagonist, but were not affected by 9-AC and niflumic acid, chloride channel blockers. GABA also produced significant membrane depolarization (19mV, n=28). As in the case of the Cl- currents, the GABA-induced depolarizations were largely blocked by bicuculline(10micrometer, n=6), but not by DIDS(50micrometer, n=4), another chloride channel blocker. CONCLUSION: The data suggest that GABAergic roles may be due to it's activation of excitatory GABAA receptors, which are expressed in MPG neurons.

Identification of ATP-sensitive K+ Conductances in Male Rat Major Pelvic Ganglion Neurons

Major pelvic ganglia (MPG) neurons are classified into sympathetic and parasympathetic neurons according to the electrophysiological properties; membrane capacitance (Cm), expression of T-type Ca2+ channels, and the firing patterns during depolarization. In the present study, function and molecular expression of ATP-sensitive K+ (K(ATP)) channels was investigated in MPG neurons of male rats. Only in parasympathetic MPG neurons showing phasic firing patterns, hyperpolarizing changes were elicited by the application of diazoxide, an activator of K(ATP) channels. Glibenclamide (10microM), a K(ATP) channel blocker, completely abolished the diazoxide-induced hyperpolarization. Diazoxide increased inward currents at high K+ (90 mM) external solution, which was also blocked by glibenclamide. The metabolic inhibition by the treatment with mitochondrial respiratory chain inhibitors (rotenone and antimycin) hyperpolarized the resting membrane potential of parasympathetic neurons, which was not observed in sympathetic neurons. The hyperpolarizing response to metabolic inhibition was partially blocked by glibenclamide. RT-PCR analysis revealed that MPG neurons mainly expressed the K(ATP) channel subunits of Kir6.2 and SUR1. Our results suggest that MPG neurons have K(ATP) channels, mainly formed by Kir6.2 and SUR1, with phenotype-specificity, and that the conductance through this channel in parasympathetic neurons may contribute to the changes in excitability during hypoxia and/or metabolic inhibition.