Pertussis toxin‐sensitive G_i‐proteins and intracellular calcium sensitivity of vasoconstriction in the intact rat tail artery

Abstract

1 We studied the involvement of pertussis toxin (PTX)-sensitive G-proteins in the sensitivity of arterial constriction to intracellular calcium ([Ca 2+ ] i ) mobilization. 2 Vasoconstriction was measured in vitro in perfused, de-endothelialized rat tail arteries loaed with the calcium-sensitive dye, fura-2 and treated or not with PTX (30 ± 1000 ng ml 71 ). Arteries were stimulated with noradrenaline (NA, 0.1 ± 100 mM) or KCl (15 ± 120 mM). 3 KCl elicited a smaller vasoconstrictor response (E max =94+8 mmHg) than NA (E max =198+9 mmHg) although [Ca 2+ ] i mobilization was similar (E max =123+8 and 135+7 nM for KCl and NA, respectively). PTX (1000 ng ml 71 ) had no e ect on [Ca 2+ ] i mobilization but lowered NA-(but not KCl-) induced vasoconstriction (E max =118+7 mmHg). 4 G i/o -proteins were revealed by immunoblotting with anti-G ia and anti-G oa antibodies in membranes prepared from de-endothelialized tail arteries. [a 32 P]-ADP-ribosylation of G-proteins by PTX (1000 ng ml 71 ) was demonstrated in the intact rat tail artery (pixels in the absence of PTX: 3150, presence: 25053). 5 In conclusion, we suggest that smooth muscle cells possess a PTX-sensitive G i -protein-mediated intracellular pathway which ampli®es [Ca 2+ ] i sensitivity of contraction in the presence of agonists such as NA.

Paradoxical Behavior of Neuromedin U in Isolated Smooth Muscle Cells and Intact Tissue

“…i and inhibit forskolin-stimulated cAMP accumulation with similarly high potency via G␣ q and G␣ i , respectively (Brighton et al, 2004b). Although we have not explored the signaling consequences of G␣ I coupling in smooth muscle cells, previous studies have suggested that G␣ i can contribute significantly to agonist-mediated contractile events (Spitzbarth-Régrigny et al, 2000), and this would be consistent with offsetting cAMP-mediated relaxation. Although there are clearly exceptions, peripheral tissue generally expresses NmU-R1, whereas NmU-R2 is generally expressed within the central nervous system, suggesting that the NmU receptor involved here is likely to be NmU-R1 (see Brighton et al, 2004a).…”

Melatonin counteracts the loss of agonist‐evoked contraction of aortic rings induced by incubation

“…NO‐mediated activation of phosphorylated myosin light chain (P‐MLC) phosphatase may be involved [27]. In another experiment, we have shown, using pertussis toxin, a G‐protein inhibitor, and tautomycine, a phosphatase inhibitor, that amplification of agonist‐evoked contraction via inhibition of P‐MLC phosphatase is a membrane receptor‐mediated event [28]. This reinforces the argument that a receptor‐mediated event is involved in the effects of melatonin observed in the present paper.…”

Cerebrovascular Gi Proteins Protect Against Brain Hypoperfusion and Collateral Failure in Cerebral Ischemia

“…On the other hand, the significance of singular knockouts is limited by overlapping functions of these isoforms. Pertussis toxin (PTX) has been used to study G i protein signaling in the cardiovascular system [7,8], but the effects on cerebrovasculature have not yet been elucidated.…”

“…In vivo, PTX irreversibly and with high specificity blocks G i -linked GPCR signal transduction -hereafter referred to as non-cerebral G i PCR KO -by catalyzing covalent modification of a C-terminal cysteine residue of cellular Gα i isoforms [9][10][11]. In arteries and microvessels, PTX inhibits endothelium-dependent relaxation to certain agonists such as β-adrenergic ligands, angiotensin, serotonin, or relaxins and is therefore useful for the study of vasculopathies [2,7,8,[12][13][14][15][16][17][18]. We have previously demonstrated that PTX, administered in a single peritoneal injection, does not cross the blood-brain barrier (BBB) and does not modify G i PCR signaling in neurons [19].…”