Journal List > Korean J Urol > v.47(5) > 1069901

Kim, Cha, Park, Song, Park, Song, and Kong: Modulatory Role of Adenylyl Cyclase and Protein Kinase A (PKA) in 5-hydroxytriptamine3 Induced Intracellular Calcium Increase in Parasympathetic Neurons of Rat Major Pelvic Ganglia

Abstract

Purpose

Serotonin has effects on the bladder contraction or urethral sphincter tone. Different subtypes of 5-hydroxytriptamine (5-HT) receptors appear to mediate the effects of serotonin on voiding. 5-HT1 and 5-HT2, metamorphic receptors, are examined well. However 5-HT3, ionotrophic receptors, are not examined well. Pelvic ganglia provide the majority of the innervation of the lower urinary tract. Major pelvic ganglia (MPG) in rats are autonomic ganglia, containing both sympathetic and parasympathetic neurons related with voiding. We examined the modulatory role of adenylyl cyclase (AC) and protein kinase A (PKA) in 5-HT3 induced intra cellular calcium increase in rat MPG.

Materials and Methods

The regulatory effects by AC and PKA were investigated in a single neuron of male rat major pelvic ganglia using patch clamp and fluorescence Ca2+ measurement techniques.

Results

Inward currents were induced by 5-HT (10 µM) at only parasympathetic neurons of MPG. MDL7222 (10-6M), selective 5-HT3 receptor antagonists, completely abolished the 5-HT induced inward currents. 5-HT (10 µM) induced intracelluar increases of calcium. These increases were blocked by an AC inhibitor SQ22536 (2×10-5M) and myristoylated PKA inhibitor (10-7M). Furthermore, foskolin (10-6M), AC activator, augmented the 5-HT induced intracellular calcium increase.

Conclusions

The activation of AC/PKA-dependent pathway could enhance 5-HT3 induced intacellular calcium increase in parasympathetic neurons of rat MPG and these is helpful for the better understanding the mechanisms of the bladder contraction or urethral sphincter tone.

Figures and Tables

Fig. 1
Effects of 5-HT3 blocker on the inward currents induced by the treatment of 10 µM 5-HT in the parasympathetic MPG neurons held at -80 mV. (A, B) The inward currents are blocked by 1 uM MDL7222, 5-HT3 blocker. 5-HT: 5-hydroxytryptamine, MPG: major pelvic ganglia.
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Fig. 2
Effects of 5-HT3 blocker on the intracellular increase of calcium induced by 10uM 5-HT. Intracelluar increases of calcium induced by the treatment of 10uM 5-HT are blocked by the treatment of 1uM MDL7222, 5-HT3 blocker, in parasympathetic MPG neurons. 5-HT: 5-hydroxytryptamine, MPG: major pelvic ganglia.
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Fig. 3
Intracellular change of calcium ion concentration induced by 5-HT in current or voltage clamp mode in parasympathetic MPG neurons. (A) Changes of intracellular calcium ion concentration and membrane potential induced by the treatment of 10uM 5-HT in current clamp mode (I=0) are recorded simultaneously. (B) Changes of intracellular calcium ion concentration and membrane currents induced by the treatment of 10uM 5-HT in current clamp mode (holding potential=-80mV) are recorded simultaneously. 5-HT: 5-hydroxytryptamine, MPG: major pelvic ganglia.
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Fig. 4
Effects of the adenylate cyclase activator on the increase of intracellular calcium induced by 5-HT in parasympathetic MPG neurons. (A, B) Intracellular calcium increase induced by 10uM 5-HT is augmented by the treatment of 1uM foskolin, adenylate cyclase activator. FSK: foskolin, Data are expressed mean±SE. *p<0.01. 5-HT: 5-hydroxytryptamine, MPG: major pelvic ganglia.
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Fig. 5
Effects of AC or PKA on the increase of intracellular calcium induced by 5-HT in parasympathetic MPG neurons. (A, B) Intracellular calcium increase induced by 10uM 5-HT is inhibited by the treatment of 200uM SQ22536, AC inhibitor and recovered without treatment. (C, D) Intracellular calcium increase induced by 10uM 5-HT is inhibited by the treatment of 100nM myristolylated PKA inhibitor and recovered without treatment. Data are expressed mean±SE. AC: adenylate cyclase, PKA: protein kinase A, WO: without treatment of SQ22536. *p<0.01, p<0.05. 5-HT: 5-hydroxytryptamine, MPG: major pelvic ganglia.
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