M(3) Subtype of Muscarinic Receptors Mediate Ca(2+) Release from Intracellular Stores in Rat Prostate Neuroendocrine Cells

Our previous studies document the expression of adrenoceptors and purinoceptors in the rat prostate neuroendocrine cells (RPNECs). However, a direct investigation of the receptors for acetylcholine (ACh) is still lacking in the prostate neuroendocrine cells. RPNECs were freshly isolated from the ventral lobes of rat prostate by using collagenase. Effects of ACh and various muscarinic antagonists on the intracellular Ca(2+) concentration ([Ca(2+)](c)) were investigated by using the fura-2 spectrofluorimetry. Single-cell RT-PCR analysis was applied to identify the transcripts for the muscarinic receptor subtypes. ACh (5 µM) induced a sharp transient increase in the [Ca(2+)](c) of RPNECs, which was independent of the extracellular Ca(2+). In the same RPNECs, high KCl (60 mM), phenylephrine (5 µM), UTP (P2Y(1/2) agonist, 50 µM), and α,β-meATP (P2X(1/3) agonist, 0.5 µM) also increased the [Ca(2+)](c). The ACh-induced [Ca(2+)](c) change (Δ[Ca(2+)](c)) was blocked by atropine or by para-fluorohexahydrosiladifenidol (M(3) antagonist, 0.3 µM), but not by telenzepine (M(1) antagonist, 1 µM) and himbacine (M(2) and M(4) antagonist, 1 µM). The single-cell RT-PCR demonstrated the selective expression of mRNAs for M(3) in RPNECs. In summary, RPNECs express M(3) muscarinic receptors that are linked to the release of Ca(2+) from intracellular stores. The Ca(2+) signals of RPNECs might mediate the parasympathetic regulation of prostate gland.