Receptor-specific contributions of caveolae, PKC, and Src tyrosine kinase to serotonergic and adrenergic regulation of Kv channels and vasoconstriction

Aims: Caveolae are invaginated, Omega-shaped membrane structures. They are now recognized as portals for signal transduction of multiple chemical and mechanical stimuli. Notably, the contribution of caveolae has been reported to be receptor-specific. However, details of how they differentially contribute to receptor signaling remain unclear. Main methods: Using isometric tension measurements, patch-clamping, and western blotting, we examined the contribution of caveolae and their related signaling pathways to serotonergic (5-HT2A receptor-mediated) and adrenergic (a1-adrenoceptor-mediated) signaling in rat mesenteric arteries. Key findings: Disruption of caveolae by methyl-beta-cyclodextrin effectively blocked vasoconstriction mediated by the 5-HT2A receptor (5-HT2AR), but not by the alpha 1-adrenoceptor. Caveolar disruption selectively impaired 5-HT2AR-mediated voltage-dependent K+ channel (Kv) inhibition, but not alpha 1-adrenoceptor-mediated Kv inhibition. In contrast, both serotonergic and alpha 1-adrenergic effects on vasoconstriction, as well as Kv currents, were similarly blocked by the Src tyrosine kinase inhibitor PP2. However, inhibition of protein kinase C (PKC) by either GO6976 or chelerythrine selectively attenuated the effects mediated by the alpha 1-adrenoceptor, but not by 5-HT2AR. Disruption of caveolae decreased 5-HT2AR-mediated Src phosphorylation, but not alpha 1-adrenoceptormediated Src phosphorylation. Finally, the PKC inhibitor GO6976 blocked Src phosphorylation by the alpha 1-adrenoceptor, but not by 5-HT2AR. Significance: 5-HT2AR-mediated Kv inhibition and vasoconstriction are dependent on caveolar integrity and Src tyrosine kinase, but not on PKC. In contrast, alpha 1-adrenoceptor-mediated Kv inhibition and vasoconstriction are not dependent on caveolar integrity, but rather on PKC and Src tyrosine kinase. Caveolae-independent PKC is upstream of Src activation for alpha 1-adrenoceptor-mediated Kv inhibition and vasoconstriction.