A similar transduction pathway involving Gs and PKA has been identified in response to activation of 3-adrenoceptors in the same myocytes (Viard direct interaction between the Ca2+ channel complex and the G subunits released from the activated G protein heterotrimer (Ikeda, 1996; De Waard PI3K; accordingly, infusion of cells with purified PI3K also stimulates L-type Ca2+ channels (Viard em et al /em ., 1999). as well as a pre-treatment with wortmannin (a PI3K inhibitor) blocked the 2-adrenergic stimulation of IBa. These results suggest that the 2-adrenergic stimulation of vascular L-type Ca2+ channels involves both Gs and G subunits which exert their stimulatory effects through PKA and PI3K/PKC pathways, respectively. the cyclic AMP/protein kinase A (PKA) transduction pathway (Ruiz-Velasco values 0.05 were considered as significant. Solutions The physiological solution used to record Ba2+ currents contained (in mM): NaCl 130, KCl 5.6, MgCl2 1, BaCl2 5, glucose 11, HEPES 10, pH?7.4 with NaOH. The basic pipette solution contained (in mM): CsCl 130, EGTA 10, ATPNa2 5, GTP 0.1, MgCl2 2, HEPES, 10 pH?7.3, with CsOH. G proteins were stored in a solution containing 20?mM Tris, 1?mM EDTA, 11?mM CHAPS, and 20?mM -mercaptoethanol. At the concentration of G used in the experiments, the final concentration of detergent was 100?M CHAPS, which alone had no effects on Ba2+ current density (Viard 2-adrenoceptors activates the Gs/PKA pathway for regulation of vascular L-type Ca2+ channels. When the G pathway was eliminated by application of calphostin C or infusion with anti-Gcom antibody or ARK1 peptide, the isoprenaline-induced stimulation of L-type Ca2+ channel was entirely abolished by PKA inhibitors (H-89 and Rp 8-Br-cyclic AMPs). A similar transduction pathway involving Gs and PKA has been identified in response to activation of 3-adrenoceptors in the same myocytes (Viard direct interaction between the Ca2+ channel complex Embramine Embramine and the G subunits released from the activated G protein heterotrimer (Ikeda, 1996; De Waard PI3K; accordingly, infusion of cells with purified PI3K also stimulates L-type Ca2+ channels (Viard em et al /em ., 1999). It is not yet clear what specific combination of G protein is coupled to Gs since the subunit composition of the Gs proteins that interact with the 2- and 3-adrenoceptors has not been identified. Different combinations of and subunits (except 11) have been reported to Embramine have similar actions on various effectors (Dolphin, 1998). However, recent data have shown that activation of mitogen-activated protein kinase/extracellular signal-regulated kinase and inhibition of adenylyl cyclases V and VI appear to be G isoform specific (G1 being more efficient than G5; Zhang em et al /em ., 1996; Bayewitch em et al /em ., 1998). Recombinant mammalian G1?C?32 complexes stimulate PI3K with similar potency and efficacity whereas G52 is not effective and appears to be unable to stimulate L-type Ca2+ channels in vascular myocytes, suggesting that signalling specificity may be encoded in the direct protein?C?protein interaction between G and PI3K (Maier em et al /em ., 2000). Obviously, selective protein?C?protein interactions represent the first step in signalling specificity and may be a possible explanation for the absence of G-activated pathway during 3-adrenoceptors activation (Viard em et al /em ., 1999). It can be postulated that the G dimers coupled to Gs may be different in relation to the existence of two forms of Gs, as previously suggested (Chaudhry & Granneman, 1991; Chaudhry em et al /em ., 1994). Additional factors such as cell compartmentation, spatial and temporal expression of transduction components may be also involved in signalling specificity, but they remain to be studied in more detail. Although three 1?C?3-adrenergic subtypes have been identified in portal vein myocytes by RT?C?PCR, we found that only 2- and 3-adrenoceptors stimulated L-type Ca2+ channels. Mixed populations of -adrenergic Rabbit polyclonal to NUDT6 receptor subtypes have been previously reported in vascular and visceral smooth muscles, but predominant roles for 2- and 3-adrenoceptors in inducing relaxation are generally demonstrated (De Boer em et al /em ., 1993; Satake em et al /em ., 1996; Yamazaki em et al /em ., 1998; Roberts em et al /em ., 1999). Existence of several -adrenoceptor subtypes producing the same physiological effect raises the question of the role of these receptors. Although a redundant function of these receptors cannot be discarded, specific modulations of -adrenoceptor subtype have been previously described. For example, 3-adrenoceptors, unlike 1- and 2-subtypes, lack regulatory phosphorylation sites for G protein receptor kinases (Liggett em et al /em ., 1993) and could be relatively resistant to agonist-induced desensitization. Thus, the function of 3-adrenoceptors may become predominant after desensitization of the 2-adrenoceptors when the sympathetic nervous system is highly stimulated. The mechanisms by which activation of -adrenoceptors induce relaxation in smooth muscles are not fully understood although several hypotheses have been proposed including Ca2+-dependent and Ca2+-independent mechanisms..
A similar transduction pathway involving Gs and PKA has been identified in response to activation of 3-adrenoceptors in the same myocytes (Viard direct interaction between the Ca2+ channel complex and the G subunits released from the activated G protein heterotrimer (Ikeda, 1996; De Waard PI3K; accordingly, infusion of cells with purified PI3K also stimulates L-type Ca2+ channels (Viard em et al /em
