CALCITONIN-GENE-RELATED PEPTIDE STIMULATES A POSITIVE CONTRACTILE RESPONSE IN RAT VENTRICULAR CARDIOMYOCYTES

Calcitonin gene-related peptide (CGRP) elicits marked positive inotropic and chronotropic actions in the atria of several mammals. The second-messenger substance cyclic AMP and activation of L-type calcium channels have been implicated in these actions, but CGRP failed consistently to stimulate a contractile response in ventricular tissue obtained from various mammals. We assessed the actions of CGRP using isolated ventricular cardiomyocytes obtained from adult rats. Maximum changes in cell length (dL) of isolated cardiomyocytes during electrically stimulated (0.5 Hz) contractions were determined with adenosine deaminase (2.5 U/ml). In these conditions, CGRP produced a potent concentration-dependent positive contractile response that became maximal 4 min after initial stimulation. CGRP increased amplitude of cellular contractions maximally at a 1-nM concentration to a value 21.4% greater than that obtained without peptide. The EC(50) value for the response was 31 pM. At concentrations greater than 1 nM, amplitude of the cellular contractile response decreased rapidly. The CGRP(2)-selective agonist, [cys ACM(2,7)] CGRP, increased the amplitude of cellular contractions maximally at 500 nM to a value 19.8% greater than that obtained without peptide. EC(50) for this response was 6 nM. Salmon calcitonin (less than or equal to 100 nM) did not elicit a significant contractile response. The fragment, CGRP(8-37), a selective antagonist at the CGRP(1) receptor subtype, while devoid of agonist activity, was a potent competitive antagonist of the positive contractile action of CGRP (pA(2) value = 7.95). CGRP, present at maximally effective concentration (1 nM), when combined with isoprenaline ISO 100 pM-1 mu M, elicited a greater increase in contractile amplitude than that elicited by ISO 100 pM-1 mu M without CGRP. CGRP 1 nM combined with low concentrations of extracellular calcium ion less than or equal to 4 mM produced a greater increase in contractile amplitude than that elicited by calcium ion less than or equal to 4 mM without CGRP, but this additive effect was abolished in the presence of higher concentrations of extracellular calcium ion (>4 mM). The cyclic AMP antagonist, Rp-cyclic AMPS (less than or equal to 200 CGM), did not inhibit the contractile response to CGRP 1 nM, but inhibited the contractile responses to ISO 100 nM and secretin 20 nM significantly and in a concentration-dependent manner. Diltiazem less than or equal to 1 mu M, a selective antagonist of L-type calcium channels, also failed to inhibit the contractile response to CGRP 1 nM but inhibited the contractile responses to ISO 100 nM and secretin 20 nM significantly and in a concentration-dependent manner. With phosphodiesterase (PDE) inhibitor, isobutyl methylxanthine (IBMX 1 mM) and adenosine deaminase 2.5 U/ml added, CGRP less than or equal to 10 mu M and salmon calcitonin less than or equal to 10 mu M failed to stimulate production of cyclic AMP in suspensions of viable cardiomyocytes, whereas secretin, included as a positive control, produced maximal stimulation four times greater than that obtained without peptide. CGRP less than or equal to 10 mu M did not stimulate production of cyclic GMP in suspensions of viable cardiomyoctyes, whereas sodium nitroprusside (SNP), used as a positive control, stimulated production maximally to a value 2.75 times greater than that obtained without agonist. These data indicate that CGRP exerts a potent positive contractile response in rat ventricular cardiomyocytes at concentrations compatible with those circulating in rat plasma. The response is mediated by receptors of the CGRP, subtype but is not coupled to a cyclic AMP- or cyclic GMP-dependent intracellular pathway and does not involve recruitment of L-type calcium channels.