Evidence that 5-HT stimulates intracellular Ca2+ signalling and activates pannexin-1 currents in type II cells of the rat carotid body.

Abstract

5-HT is a neuromodulator released from carotid body (CB) chemoreceptor (type I) cells and facilitates the sensory discharge following chronic intermittent hypoxia (CIH). In the present study, we show that, in addition to type I cells, adjacent glial-like type II cells express functional, ketanserin-sensitive 5-HT2 receptors, and their stimulation increases cytoplasmic Ca2+ derived from intracellular stores. In type II cells, 5-HT activated a ketanserin-sensitive inward current (I5-HT ) that was similar to that (IUTP ) activated by the P2Y2R agonist, UTP. As previously shown for IUTP , I5-HT was inhibited by BAPTA-AM and carbenoxolone (5μm), a putative blocker of ATP-permeable pannexin (Panx)-1 channels; IUTP was reversibly inhibited by the specific Panx-1 mimetic peptide channel blocker, 10 Panx peptide. Paracrine stimulation of type II cells by 5-HT, leading to ATP release via Panx-1 channels, may contribute to CB excitability, especially in pathophysiological conditions associated with CIH (e.g. obstructive sleep apnoea). Carotid body (CB) chemoreceptor (type I) cells can synthesize and release 5-HT and increased autocrine-paracrine 5-HT2 receptor signalling contributes to sensory long-term facilitation during chronic intermittent hypoxia (CIH). However, recent studies suggest that adjacent glial-like type II cells can respond to CB paracrine signals by elevating intracellular calcium (Δ[Ca2+ ]i ) and activating carbenoxolone-sensitive, ATP-permeable, pannexin (Panx)-1-like channels. In the present study, using dissociated rat CB cultures, we found that 5-HT induced Δ[Ca2+ ]i responses in a subpopulation of type I cells, as well as in most (∼67%) type II cells identified by their sensitivity to the P2Y2 receptor agonist, UTP. The 5-HT-induced Ca2+ response in type II cells was dose-dependent (EC50 ∼183nm) and largely inhibited by the 5-HT2A receptor blocker, ketanserin (1μm), and also arose mainly from intracellular stores. 5-HT also activated an inward current (I5-HT ) in type II cells (EC50 ∼200nm) that was reversibly inhibited by ketanserin (1-10nm), the Ca2+ chelator BAPTA-AM (5μm), and low concentrations of carbenoxolone (5μm), a putative Panx-1 channel blocker. I5-HT reversed direction at approximately -11mV and was indistinguishable from the UTP-activated current (IUTP ). Consistent with a role for Panx-1 channels, IUTP was reversibly inhibited by the specific Panx-1 mimetic peptide blocker 10 Panx (100μm), although not by its scrambled control peptide (sc Panx). Because ATP is an excitatory CB neurotransmitter, it is possible that the contribution of enhanced 5-HT signalling to the increased sensory discharge during CIH may occur, in part, by a boosting of ATP release from type II cells via Panx-1 channels.