Receptor-mediated shifts in cGMP and cAMP levels in neuroblastoma cells.

Abstract

In this report the effects of activators of inhibitory muscarinic acetylcholine receptors, adenosine receptors, and prostaglandin El (PGEj) receptors (1-5) on 3':5'-cGMP and 3':5'-cAMP levels of neuroblastoma cells are described. Muscarinic acetylcholine receptors frequently mediate slow, prolonged responses which may be excitatory or inhibitory; in contrast to the nicotinic acetylcholine receptor, which in higher organisms mediates fast excitatory responses. Activation of the inhibitory or excitatory muscarinic acetylcholine receptors of cardiac (6, 7) and smooth muscle (7) results in an increase in cGMP concentration. We find that cGMP levels of neuroblastoma Cell Culture. The following cell lines were used: mouse neuroblastoma C-1300 clones NlE-115 and N18 (8), mouse neuroblastoma X mouse L cell hybrid clone NL1F (9), and rat glioma clone C6BU-1 (10). Neuroblastoma cells were grown in 100 mm petri dishes (Falcon, 55 cm2 surface area) in 15 ml of 90% DMEM-10% fetal-bovine serum (340 mOsm/kg) in a humidified atmosphere of 10% C02-90% air at 370. Hybrid cells were grown in the same medium supplemented with 0.1 mM hypoxanthine, 1 ,uM aminopterin, and 16 ,gM thymidine (HAT). Cells usually were grown to confluency (1 to 3 X 105 cells, 0.1 to 0. Purification of cGMP and cAMP. A rapid method was devised for separating cGMP from cAMP and removing trichloroacetic acid and endogenous interfering compounds from cyclic nucleotide fractions. The supernatant solution of the deproteinized sample in 5% trichloroacetic acid was applied to an ion exchange column (0.7 X 8 cm) of

Evidence for involvement of cyclic nucleotides in intrinsic factor secretion by isolated rabbit gastric mucosa.

“…In vitro effects of humoral agents on cyclic GMP (cGMP) levels are often very early and short-lived (16)(17)(18). To measure levels of cyclic nucleotides at very short incubation periods (1-2 min), we modified the culture technique.…”

Neurotransmitter receptors mediate cyclic GMP formation by involvement of arachidonic acid and lipoxygenase.

“…Murine neuroblastoma clone NlE-115 cells possess muscarinic and histamine H1 receptors that mediate cyclic GMP formation (3-6). These cells, derived from a neuroblast, display many of the properties of normal differentiated neurons (7) and, as such, have proven to be an ideal model system for the study of the mechanisms of neurotransmitter interactions with nerve cells, including receptor activation, desensitization, and psychotherapeutic drug effects (3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14).…”

Effect of acetylcholine on the norepinephrine-induced positive chronotropy and increase in cyclic nucleotides of isolated rabbit sinoatrial node.

“…Such muscarinic receptor-linked changes in the cyclic GMP levels have been demonstrated in the heart (George et al, 1970;Watanabe and Besch, 1975;Gardner and Allen, 1976), smooth muscles (Schultz et al, 1973), secretory systems (Smith and Ignarro, 1975;Haymovits and Scheele, 1976), cultured neurons (Matsuzawa and Nirenberg, 1975), and brain slices (Ferrendelli et al, 1970;Hanley and Iversen, 1978). Our results are similar to such findings with respect to the transient accumulation of cyclic GMP in the SA node after an application of acetylcholine.…”