USE OF D-MYO INOSITOL 1,2,6-TRISPHOSPHATE TO INHIBIT CONTRACTILE ACTIVITY IN RAT VENTRICULAR CARDIOMYOCYTES INDUCED BY NEUROPEPTIDE-Y AND OTHER CARDIOACTIVE PEPTIDES THROUGH PHOSPHOLIPASE-C

1 D-Myo inositol 1,2,6 trisphosphate (alpha-trinositol, pp56), an isom er of the second messenger substance, inositol 1,4,5 trisphosphate, ha s an interesting pharmacological profile that includes antagonism of a number of neuropeptide Y (NPY)-mediated cellular processes. The abili ty of pp56 to inhibit selectively the myocardial contraction mediated by NPY in relation to the responses to other cardioactive peptides, in cluding endothelin-1, calcitonin gene-related peptide (CGRP), secretin and vasoactive intestinal peptide (VIP), was assessed. In order to in vestigate the possible interaction of pp56 with mechanisms of inositol phosphate signalling generated in heart muscle cells by activation of the beta-isoenzyme of phospholipase C (PLC beta), noradrenaline was u sed as a positive control, and isoprenaline and forskolin were include d as negative controls. 2 Ventricular cardiomyocytes, isolated from th e hearts of adult rats, were stimulated to contract at 0.5 Hz in the p resence of calcium ion (2 mM). The concentrations of agonists used wer e in the region of their maximally effective concentrations for myocyt e contraction and the concentration of pp56 was in the range of 1-100 mu M. Contractile activity was monitored by video microscopy and maxim um shortening determined by image analysis. 3 In the absence of agonis t, contractile amplitudes following 20 min preincubation with pp56 wer e not different from that observed in the absence of pp56. Pp56 (1-100 mu M) inhibited significantly the positive contractile response to no radrenaline (5 mu M) in the presence of propranolol (500 nM), such tha t the response was almost completely attenuated at the highest concent ration of the inhibitor. Pp56 did not inhibit the positive contractile responses to forskolin (40 mu M) or isoprenaline (100 nM). 4 NPY alon e does not influence the basal level of contraction of cardiomyocytes, but can attenuate isoprenaline-stimulated contraction and can increas e contractile amplitude from basal when the transient outward current is blocked with 4-aminopyridine. In the presence of isoprenaline (100 nM), the negative response to NPY (100 nM) was attenuated significantl y by pp56 (1-100 mu M). With 4-aminopyridine, the positive contractile response to NPY (200 nM) was decreased by pp56, although this was not statistically significant. 5 Pp56 inhibited the positive contractile responses to CGRP (1 nM) and endothelin-1 (20 nM) completely, but did not affect the responses to secretin (20 nM) or VIP (20 nM). 6 In conc lusion, these data challenge the previously obtained selectivity of pp 56 as an antagonist of NPY-mediated cellular processes, since response s to CGRP and endothelin-1 were at least equally sensitive. Furthermor e, as pp56 discriminated clearly in its inhibition of responses to alp ha-adrenoceptor by comparison with beta-adrenoceptor/adenylate cyclase stimulation, it appears that pp56 may be a useful pharmacological age nt with which to distinguish between PLC beta-dependent and PLC beta-i ndependent coupling mechanisms. On this basis, further evidence has be en obtained that, in rat cardiomyocytes, the contractile responses to NPY, CGRP and endothelin-1 are attributable to the activation of PLC b eta-dependent pathways, whereas the responses to secretin and VIP are mediated by PLC beta-independent pathways.