calcimycin and inositol-1-4-bis(phosphate)

Synaptosomes have been isolated from rat cerebral cortex and labelled in vitro with [32P]orthophosphate and myo-[2-3H]inositol. Subsequent addition of the Ca2+ ionophore A23187 in the presence of 2 mM extrasynaptosomal Ca2+ raised intrasynaptosomal free [Ca2+] to greater than 2 microM from a resting level of 200 nM and led to rapid breakdown of polyphosphoinositides. This was accompanied by a small increase in the level of inositol monophosphate, greatly enhanced accumulation in inositol bisphosphate, but no detectable increase in inositol trisphosphate. Depolarising (25 mM) extrasynaptosomal K+ produced a smaller increase in intrasynaptosomal free [Ca2+] (to around 400 nM) and a proportional increase in inositol bisphosphate radioactivity. Carbachol (1 mM) alone elicited only limited polyphosphoinositide breakdown and inositol mono- and bisphosphate formation, but this was greatly increased in the presence of 25 mM K+. The effect of carbachol in the presence of depolarising K+ was time- and dose-dependent and was antagonised by atropine (10 microM). There was no detectable accumulation of inositol trisphosphate in the presence of carbachol, K+, or carbachol plus K+, even after short (30 s.) incubations. The lack of inositol trisphosphate accumulation does not appear to result from rapid formation of inositol tetrakisphosphate or from enhanced breakdown of the trisphosphate in synaptosomes.

Topics: Animals; Calcimycin; Carbachol; Inositol Phosphates; Male; Phosphatidylinositol Phosphates; Phosphatidylinositols; Potassium; Rats; Rats, Inbred Strains; Sugar Phosphates; Synaptosomes

The vasoactive intestinal peptide (VIP) induces a concentration-dependent secretion of newly synthesized (3H labeled) proteins from lacrimal gland fragments. Maximal secretory response is approximately 20% of total labeled proteins secreted for a 40-min stimulation and half-maximal secretory response is obtained at 3.8 +/- 0.2 nM VIP. The cholinergic (muscarinic) and VIPergic stimulations synergistically interact in eliciting newly synthesized protein secretion. Carbachol (0.3 microM) and the phorbol ester PMA (1 microM) potentiate the secretory response to VIP (10 nM), forskolin (3 microM), and dibutyryl adenosine 3',5'-cyclic monophosphate (DBcAMP) (0.5 mM) both in the absence and presence of 2.5 mM extracellular calcium. The calcium ionophore A23187 (1 microM) potentiates the cAMP-dependent responses only in the presence of extracellular calcium. We propose that newly synthesized protein secretion from rat lacrimal glands is controlled by two systems interacting synergistically at a step distal to the production of intracellular second messengers. The potentiating effect of agonists acting through the calcium-dependent pathway on the cAMP-dependent secretory response may involve both calcium and diacylglycerol.

Topics: Animals; Bucladesine; Calcimycin; Carbachol; Colforsin; Inositol 1,4,5-Trisphosphate; Inositol Phosphates; Lacrimal Apparatus; Male; Papaverine; Proteins; Rats; Rats, Inbred Strains; Tetradecanoylphorbol Acetate; Vasoactive Intestinal Peptide

In the guinea pig myometrium prelabelled with myo-[2-3H]inositol, carbachol and oxytocin enhanced a concentration-dependent and rapid release of IP3 which preceded that of IP2 and IP1. The specific receptor-mediated phospholipase C activation degrading PIP2 to IP3 did not require the presence of extracellular Ca2+. The ionophore A23187 as well as K+ depolarization failed to increase inositol phosphate accumulation. It is proposed that IP3 could have a role in the contraction of uterine smooth muscle elicited by the activation of muscarinic as well as of oxytocin receptors.

Topics: Animals; Calcimycin; Calcium; Carbachol; Chlorides; Female; Guinea Pigs; Inositol 1,4,5-Trisphosphate; Inositol Phosphates; Kinetics; Lithium; Lithium Chloride; Myometrium; Oxytocin; Receptors, Angiotensin; Receptors, Muscarinic; Receptors, Oxytocin; Sugar Phosphates; Type C Phospholipases