Mechanisms of cGMP-dependent mesangial-cell relaxation: a role for myosin light-chain phosphatase activation.

Although the cGMP-dependent relaxation of contractile cells seems to depend on the ability of the cyclic nucleotide to interfere with intracellular calcium, this does not appear to be the only mechanism involved. The present experiments were designed to analyse alternative mechanisms, trying to test the hypothesis that cGMP could relax rat mesangial cells by activating myosin light-chain phosphatase (MLC-PP), with the subsequent dephosphorylation of myosin light chain (MLC). The effect of a cGMP analogue, dibutyryl cGMP (dbcGMP), on angiotensin II-(AII) and PMA-induced MLC phosphorylation (MLCP) was tested, in the presence of calyculin A (CA), an inhibitor of MLC-PP. MLCP was measured, after cell labelling with (32)P, by immunoprecipitation. dbcGMP prevented the increased MLCP induced by AII or PMA, and this inhibition was blocked by CA. dbcGMP also increased the MLC dephosphorylation observed in cells incubated with AII and in which MLC kinase and protein kinase C activities were blocked. The AII-elicited increased intracellular calcium concentration was only partially inhibited by dbcGMP. These results suggest that the cGMP-induced mesangial-cell relaxation could be due, at least partially, to the stimulation of MLC-PP.

Dibutyryl cyclic guanosine monophosphate elevates bovine endothelial cell thromboxane production without affecting prostacyclin metabolism.

Cloned bovine aortic endothelial cells (BAEC) were grown to confluence then treated for 24 hours with dibutyryl cyclic AMP (dbcAMP) or dibutyryl cyclic GMP (dbcGMP) in serum free medium. Submillimolar concentrations of dbcGMP caused a significant enhancement of thromboxane (TXB2) synthesis in washed cells exposed to arachidonate. DbcAMP had no effect on the production of either metabolite. TXB2 synthesis was inhibited by 3 micrograms/ml cycloheximide, whether or not the cells were pretreated with dbcGMP. Prostacyclin production was inhibited to a much lesser extent by cycloheximide. We conclude that dbcGMP elevates TXB2 production by increasing the amount of thromboxane synthetase available, and that this effect is inhibited by cycloheximide. Data are described which suggest that dbcGMP increases the degradation rate of TXB2 by BAEC, so that the observed increase in TXB2 may be an underestimate of the true effects of dbcGMP on TXB2 production.

Opposing actions of dibutyryl cyclic AMP and GMP on temperature in conscious guinea-pigs.

Intracerebroventricular administration of dibutyryl cyclic AMP (Db-cAMP), induced hyperthermia in guinea-pigs which was not mediated through prostaglandins (PG) or norepinephrine since a prostaglandin synthesis inhibitor, indomethacin, and an alpha-adrenergic receptor blocking agent, phenoxybenzamine did not antagonize the hyperthermia. In contrast, the hyperthermic response to dibutyryl cyclic AMP was attenuated by central administration of a beta-adrenergic receptor antagonist, sotalol, indicating that cyclic AMP may be involved, through beta-adrenergic receptors, in the central regulation of heat production/conservation. Central administration of dibutyryl cyclic GMP (Db-cGMP) produced hypothermia which was not mediated via histamine H1- or H2-receptors and serotonin since the H1-receptor antagonist, mepyramine, the H2-receptor antagonist, cimetidine, and the serotonin antagonist, methysergide, had no antagonistic effects. The antagonism of hypothermia induced by dibutyryl cyclic GMP and acetylcholine + physostigmine, by central administration of a cholinergic muscarinic receptor antagonist, atropine, and not by a cholinergic nicotinic receptor antagonist, d-tubocurarine, suggests that cholinoceptive neurons and endogenous cyclic GMP may regulate heat loss through cholinergic muscarinic receptors. These results support a regulatory role in thermoregulation provided by a balance between opposing actions of cyclic AMP and cyclic GMP in guinea-pigs.