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Abstract: It has been postulated that in blood vessels under α1-related stimulation, the endothelial intracellular calcium concentration ([Ca2+] i ) increases, which is necessary to induce nitric oxide synthesis, is the result of an increase in vascular smooth muscle, which subsequently, flows into the endothelial cells through gap junctions and it is not the result of a direct adrenergic stimulation of endothelial receptors. Others, however, postulate that endothelial α1D-adrenoceptors, have a direct effect on nitric oxide synthesis. In order to clarify this phenomena, in this work we analyzed the presence of α1receptor subtypes and their functional association with nitric oxide synthesis in rat coronary microvascular endothelial cells in culture, with pharmacological, immunological and reverse transcriptase polymerase chain reaction approaches. Our results show the presence and functional coupling with nitric oxide synthesis of α1A and α1D-adrenoceptor subtypes. α1B-adrenoceptor subtype is not coupled with nitric oxide production. [Copyright &y& Elsevier]

It has been postulated that in blood vessels under alpha(1)-related stimulation, the endothelial intracellular calcium concentration ([Ca(2+)](i)) increases, which is necessary to induce nitric oxide synthesis, is the result of an increase in vascular smooth muscle, which subsequently, flows into the endothelial cells through gap junctions and it is not the result of a direct adrenergic stimulation of endothelial receptors. Others, however, postulate that endothelial alpha(1D)-adrenoceptors, have a direct effect on nitric oxide synthesis. In order to clarify this phenomena, in this work we analyzed the presence of alpha(1)receptor subtypes and their functional association with nitric oxide synthesis in rat coronary microvascular endothelial cells in culture, with pharmacological, immunological and reverse transcriptase polymerase chain reaction approaches. Our results show the presence and functional coupling with nitric oxide synthesis of alpha(1A) and alpha(1D)-adrenoceptor subtypes. alpha(1B)-adrenoceptor subtype is not coupled with nitric oxide production.

Caveolins are a family of integral membrane proteins implicated in various cell functions, including the organization and inactivation of signaling molecules of G protein-coupled receptors. We tested the ability of human caveolin scaffolding peptide-1 (CSP-1) to regulate norepinephrine- (NE) or histamine (HIS)-induced increases on intracellular calcium concentrations ([Ca(2+)]i). In cultured rat vascular smooth muscle cells (VSMC), CSP-1 inhibited in a concentration-dependent manner NE- and HIS-induced increases in [Ca(2+)]i. This effect can be explained by the fact that CSP-1 inhibited a common signaling pathway. We tested the ability of this peptide to decrease the activation of PLC-beta3 and MAPK. CSP-1 inhibited the expression of the activated form of both enzymes, suggesting a direct effect of the peptide on the signaling cascade. CSP-1 readily enters VSMC in culture, as observed when FITC-conjugated CPS-1 is added to cell culture media. Taken together, these data suggest that CSP-1 blocks the effects of NE and HIS on [Ca(2+)]i of VSMC by inhibiting the activation of PLC-beta3 and MAPK.