Evidence for protein kinase involvement in the 5‐HT‐ [Ca2+]i ‐ pannexin‐1 signalling pathway in type II glial cells of the rat carotid body

New Findings

What is the central question of this study?

The mammalian carotid body (CB) is a peripheral chemoreceptor organ involved in O₂ and CO₂/H⁺ homeostasis. Recent studies suggest that 5‐HT, released from CB receptor cells, can stimulate adjacent glial‐like type II cells, leading to a rise in intracellular Ca²⁺ ([ΔCa²⁺_i]) and activation of ATP‐permeable pannexin‐1 (Panx‐1) channels. The aim of this study was to elucidate the role of protein kinases in the 5‐HT‐[Ca²⁺]_i‐Panx‐1 signalling pathway.

What is the main finding and its importance?

Src family kinase (SKF) and PKA, acting downstream from [ΔCa²⁺_i], played central roles in 5‐HT‐mediated Panx‐1 channel activation. This provides new insight into mechanisms regulating CB excitation, especially in pathophysiological conditions.

Abstract

Chemoreceptor (type I) cells of the rodent carotid body (CB) synthesize and release several neurotransmitters/neuromodulators including 5‐hydroxytryptamine (5‐HT), implicated in enhanced CB excitation following exposure to chronic intermittent hypoxia, e.g. sleep apnea. However, recent studies suggest that 5‐HT can robustly stimulate adjacent glial‐like type II cells via ketanserin‐sensitive 5‐HT2 receptors, leading to intracellular Ca²⁺ elevation [ΔCa²⁺_i] and activation of ATP‐permeable pannexin‐1 (Panx‐1) channels. Using dissociated rat CB cultures, we investigated the role of protein kinases in the intracellular signalling pathways in type II cells. In isolated type II cells, 5‐HT activated a Panx‐1‐like inward current (I_5‐HT) that was reversibly inhibited by the Src family kinase (SFK) inhibitor PP2 (1 μM), but not by its inactive analog PP3 (1 μM). Moreover, I_5‐HT was reversibly inhibited (> 90%) by H‐89 (1 μM), a PKA blocker, whereas the PKC blocker GF109203X (2 μM) was largely ineffective. By contrast, the P2Y2R agonist UTP (100 μM) activated Panx‐1‐like currents that were reversibly inhibited (∼60%) by either H‐89 or GF109203X. Using fura‐2 spectrofluorimetry, the 5‐HT‐induced [ΔCa²⁺_i] was unaffected by PP2, H‐89, and GF109293X, suggesting the kinases acted downstream of the Ca²⁺ rise. Because intracellular Ca²⁺ chelation was previously shown to block receptor‐mediated, Panx‐1 current activation in type II cells, these data suggest that CB neuromodulators use overlapping, but not necessarily identical, signalling pathways to activate Panx‐1 channels and release of ATP, a CB excitatory neurotransmitter. In conclusion, these studies provide new mechanistic insight into 5‐HT signalling in the CB that has pathophysiological relevance.

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