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 (α‐trinositol, pp56), an isomer of the second messenger substance, inositol 1,4,5 trisphosphate, has an interesting pharmacological profile that includes antagonism of a number of neuropeptide Y (NPY)‐mediated cellular processes. The ability of pp56 to inhibit selectively the myocardial contraction mediated by NPY in relation to the responses to other cardioactive peptides, including endothelin‐1, calcitonin gene‐related peptide (CGRP), secretin and vasoactive intestinal peptide (VIP), was assessed. In order to investigate the possible interaction of pp56 with mechanisms of inositol phosphate signalling generated in heart muscle cells by activation of the β‐isoenzyme of phospholipase C (PLCβ), noradrenaline was used as a positive control, and isoprenaline and forskolin were included as negative controls. 2 Ventricular cardiomyocytes, isolated from the hearts of adult rats, were stimulated to contract at 0.5 Hz in the presence of calcium ion (2 m m ). The concentrations of agonists used were in the region of their maximally effective concentrations for myocyte contraction and the concentration of pp56 was in the range of 1–100 μ m . Contractile activity was monitored by video microscopy and maximum shortening determined by image analysis. 3 In the absence of agonist, contractile amplitudes following 20 min preincubation with pp56 were not different from that observed in the absence of pp56. Pp56 (1–100 μ m ) inhibited significantly the positive contractile response to noradrenaline (5 μ m ) in the presence of propranolol (500 n m ), such that the response was almost completely attenuated at the highest concentration of the inhibitor. Pp56 did not inhibit the positive contractile responses to forskolin (40 μ m ) or isoprenaline (100 n m ). 4 NPY alone does not influence the basal level of contraction of cardiomyocytes, but can attenuate isoprenaline‐stimulated contraction and can increase contractile amplitude from basal when the transient outward current is blocked with 4‐aminopyridine. In the presence of isoprenaline (100 n m ), the negative response to NPY (100 n m ) was attenuated significantly by pp56 (1–100 μ m ). With 4‐aminopyridine, the positive contractile response to NPY (200 n m ) was decreased by pp56, although this was not statistically significant. 5 Pp56 inhibited the positive contractile responses to CGRP (1 n m ) and endothelin‐1 (20 n m ) completely, but did not affect the responses to secretin (20  nM) or VIP (20 n m ). 6 In conclusion, these data challenge the previously obtained selectivity of pp56 as an antagonist of NPY‐mediated cellular processes, since responses to CGRP and endothelin‐1 were at least equally sensitive. Furthermore, as pp56 discriminated clearly in its inhibition of responses to α‐adrenoceptor by comparison with β‐adrenoceptor/adenylate cyclase stimulation, it appears that pp56 may be a useful pharmacological agent with which to distinguish between PLCβ‐dependent and PLCβ‐independent coupling mechanisms. On this basis, further evidence has been obtained that, in rat cardiomyocytes, the contractile responses to NPY, CGRP and endothelin‐1 are attributable to the activation of PLCβ‐dependent pathways, whereas the responses to secretin and VIP are mediated by PLCβ‐independent pathways.