1 The L6 myocyte cell line expresses high affinity receptors for calcitonin gene-related peptide (CGRP) which are coupled to activation of adenylyl cyclase. The biochemical pharmacology of these receptors has been examined by radioligand binding or adenosine 3':5'-cyclic monophosphate (cyclic AMP) accumulation. 5 The pharmacological profile of the L6 CGRP receptor suggests that it most closely resembles sites on skeletal muscle, cardiac myocytes and hepatocytes. The L6 cell line should be a stable homogeneous model system in which to study CGRP mechanisms and pharmacology.

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1 Bradykinin can release neuronal calcitonin gene-related peptide (CGRP) and adrenal medullary catecholamines, both of which could contribute to its cardiovascular effects in vivo. Therefore, in the main experiment, regional haemodynamic responses to bolus injections of bradykinin (3 nmol kg- ', i.v.) were assessed in the same chronically-instrumented, conscious, Long Evans rats in the absence and in the presence of human a-CGRP or ICI 118551, antagonists of CGRPI-receptors and P2-adrenoceptors, respectively. The selected doses of these antagonists caused specific inhibition of responses mediated by exogenous human a-CGRP and P2-adrenoceptor agonists, respectively.2 Bradykinin administered alone as an i.v. bolus had a slight pressor effect accompanied by a marked tachycardia. There were early (at about 30 s) increases in flow and conductance in the mesenteric vascular bed, and delayed (at about 90 s), but qualitatively similar, changes in the hindquarters vascular bed. There were only slight increases in flow and conductance in the renal vascular bed. 3 Human a-CGRP had no statistically significant effects on the responses to bolus doses of bradykinin. However, in the presence of ICI 118551, the pressor effect of bradykinin was significantly enhanced while its tachycardic effect was significantly suppressed. The hindquarters vasodilator effect of bradykinin was converted to a vasoconstriction and there was a slight renal vasoconstriction, but the mesenteric vasodilator effect of bradykinin was unchanged by ICI 118551. 4 In subsidiary experiments, in other animals, it was found that infusion of bradykinin (36 nmol kg-' min-') elicited a pattern of haemodynamic responses similar to that seen with bolus injections and, as in the latter case, the hindquarters hyperaemic vasodilatation was inhibited by ICI 118551. In the presence of mecamylamine (at a dose sufficient to block reflex heart rate responses to rises or falls in arterial blood pressure) bolus injection or infusion of bradykinin still elicited increases in renal, mesenteric and hindquarters blood flow. However, in additional experiments in adrenal demedullated rats (n = 4) the hindquarters hyperaemic effect of bradykinin was absent, although the mesenteric hyperaemic effect remained. 5 The results indicate that the increase in hindquarters blood flow following administration of bradykinin in vivo is largely due to activation of P2-adrenoceptors by catecholamines released subsequent to direct stimulation of the adrenal medulla by the peptide. However, the bradykinin-induced increase in mesenteric blood flow does not depend on this mechanism.

Near-u.v. and far-u.v. c.d. spectra of human alpha-calcitonin-gene-related peptide (h alpha CGRP), analogues and fragments of CGRP and amylin were recorded in aqueous solution and in trifluoroethanol (TFE)/water mixtures. All peptides contained significant amounts of alpha-helix in aqueous solution, and this amount increased on adding TFE. The helical content was unaffected by pH and salt. However, amylin contained much less helix than CGRP and the c.d. spectrum was more temperature-sensitive. A band in the near-u.v. c.d. spectrum of CGRP (but not present in the spectrum of amylin) was attributed to the disulphide bond in CGRP. The intensity of this band was pH-dependent and titrated with a pKa of 6.5, suggesting the involvement of histidine ionization.

Rat synthetic amidated islet amyloid polypeptide (IAPP) was infused into conscious Long-Evans rats chronically instrumented for the measurement of regional hemodynamics. Rat IAPP (0.25-2.5 nmol.kg-1.min-1) had dose-dependent tachycardiac and hypotensive effects. Renal blood flow increased at all dose levels in association with incremental rises in renal vascular conductances. Hindquarters blood flow and vascular conductance increased at the higher dose levels, but mesenteric blood flow fell with mean arterial blood pressure (i.e., there was no change in mesenteric vascular conductance). Concurrent infusion of 25 nmol.kg-1.min-1 human alpha-calcitonin gene-related peptide (CGRP) (8-37) abolished the hypotensive, tachycardiac, and renal and hindquarters vasodilator effects of rat IAPP, and during administration of both peptides, there was a transient renal and sustained mesenteric vasoconstriction. When the infusion of human alpha-CGRP (8-37) was stopped, the effects of the continued infusion of rat IAPP were reestablished. The results indicate that the reported ability of IAPP to induce insulin resistance cannot be due to decreased skeletal muscle blood flow. In addition, human alpha-CGRP (8-37) is an effective antagonist of the hemodynamic actions of rat IAPP. Because it has been shown previously that human alpha-CGRP (8-37) antagonizes the hemodynamic effects of human alpha-CGRP, these results, collectively, indicate that human alpha-CGRP and rat IAPP might act on the same receptor at which human alpha-CGRP (8-37) is an effective antagonist or that the latter is a nonselective antagonist of separate receptors on which human alpha-CGRP and rat IAPP act.