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1. The effects of the muscarinic agonist carbachol on phosphoinositide metabolism and its relationship to alteration of intracellular calcium were examined in SK-N-SH human neuroblastoma cells. Muscarinic receptors on these cells are coupled to phospholipase C and the myo [2-3H]-inositol phosphates resulting from receptor activation of cells labelled with [3H]-inositol accumulate rapidly. The breakdown of both inositol monophosphate (InsP1) and inositol bisphosphate (InsP2) is sensitive to lithium with inhibition of the latter only observed at higher concentrations of this ion. 2. Use of the calcium indicator dye Fura 2 revealed that carbachol stimulates a biphasic increase in intracellular calcium. 3. Carbachol was able to stimulate both [3H]-inositol phosphate production and intracellular calcium levels with respective EC50 values of 15.9 +/- 1.0 microM and 10.7 +/- 3.2 microM, indicating that no amplification occurs between these steps in the signal transduction pathway. 4. Inositol 1,4,5 trisphosphate (Ins(1,4,5)P3) released 45Ca2+ in a stereospecific and dose-related manner from intracellular stores of permeabilised cells. 5. These results suggest that this cell line may represent a useful model system to investigate receptor-mediated phosphoinositide metabolism and calcium homeostasis.

We report a case of familial porphyria cutanea tarda (FPCT) in a 7-year-old girl. The condition was exacerbated by low dose of hydroxychloroquin, but controlled by venesection. The favorable prognosis of FPCT should be further established by long-term prospective studies.

The present study describes the effect of nutrient secretagogues upon insulin release from an insulinoma derived B-cell line RINm5F. Glucose failed to stimulate insulin release, it did however increase the rate of glucose oxidation by some 16 fold. Glyceraldehyde stimulated insulin release in a concentration dependent manner, maximum stimulation occurring at 20 mmol/l. The response to glyceraldehyde was suppressed by calcium channel blockade and calmodulin inhibition. Insulin release was also stimulated by the amino acids leucine and arginine, maximum stimulation for both amino acids occurring at 15 mmol/l. The present study confirms the usefulness of this cell line as a model for the study of insulin secretion.

Acetylcholine stimulated the release of insulin from cultured HIT-T15 and RINm5F cells in the absence of glucose confirming that the insulin response to cholinergic stimulation is preserved in these two cloned B-cell lines. In addition acetylcholine potentiated glucose stimulated insulin release from HIT-T15 but not from RINm5F cells. Perifused HIT-T15 cells responded to square wave acetylcholine challenge with a monophasic release of insulin while the release profile for RINm5F cells was irregular and reduced. Acetylcholine stimulated insulin release from both cell lines was reduced by atropine but unaffected by hexamethonium confirming a mechanism of action involving muscarinic receptor activation.

The present study examines the muscarinic receptor binding characteristics of parent human neuroblastoma (SK-N-SH) and its neuroblast (SH-SY5Y) and epithelial-like (SH-EP1) clones using [3H]methylscopolamine [( 3H]NMS). Specific [3H]NMS binding to intact SK-N-SH and SH-SY5Y cells was saturable with a Kd of 0.2 nM and Bmax of 100-150 fmol/mg protein. Specific [3H]NMS binding to whole cell preparations of SH-EP 1 could not be detected. Pharmacological analysis of the binding site both in whole cells and membranes of SK-N-SH are indicative of an homogeneous receptor population possessing low affinity for the M1-selective antagonist pirenzepine. The muscarinic receptors expressed by the neuroblast clone, SH-SY5Y were further characterized and shown to have the properties of an homogeneous M3 subtype with low affinity for the M1-selective antagonist pirenzepine and the M2-cardioselective AFDX-116 but high affinity for 4-diphenylacetoxy-N-methyl piperidine methiodide (4-DAMP). In conclusion the SH-SY5Y neuroblastoma should provide an important human neuronal cell model with which to define the regulation of post-receptor events driven by a single receptor population.