Purinergic receptors coupled to intracellular Ca²⁺ signals and exocytosis in rat prostate neuroendocrine cells

Rat prostate neuroendocrine cells (RPNECs) display a variety of ion channels and exhibit α-adrenergic regulation of cytosolic Ca²⁺ concentration ([Ca²⁺]_c). In this study, purinergic regulation of [Ca²⁺]_c and exocytosis was investigated in freshly isolated single RPNECs showing chromogranin A immunoreactivity. The presence of P2X and P2Y receptors in RPNECs was verified by the transient activation of Ca²⁺-permeable cationic channels and the release of Ca²⁺ from intracellular stores by extracellular ATP, respectively. The transient inward cationic current was effectively activated by α,β-methyleneadenosine 5′-triphosphate (α,β-MeATP) and blocked by 2′,3′-O-(2,4,6-trinitrophenyl)adenosine 5′-triphosphate, suggesting the presence of a P2X₁ or P2X ₃ subtype. For the release of stored Ca²⁺, ATP and UTP were equally potent, indicating the functional expression of the P2Y₂ or P2Y₄ subtype. The mRNAs for P2X₁ and P2Y₂ were confirmed from reverse transcription-PCR analysis of RPNECs. The application of α,β-MeATP induced large and transient increases in [Ca²⁺]_c which were not attenuated by the blockers of voltage-activated Ca²⁺ channels or by depleting intracellular Ca ²⁺ stores, but were abolished by omitting extracellular Ca ²⁺. The application of UTP increased [CA²⁺]_c to 55% of the peak Δ[Ca²⁺]_c induced by α,β-MeATP. The application of α,β-MeATP induced exocytotic responses of RPNECs as monitored by carbon fiber amperometry and capacitance measurements. To our interest, the application of UTP did not induce amperometric currents, but reduced the membrane capacitance, indicating a net endocytosis. From these results, we postulate that a sharp rise in [Ca ²⁺]_c by the P2X-mediated Ca²⁺ influx is required for exocytosis, whereas the relatively slow release of stored Ca ²⁺ induces endocytosis in RPNECs.