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The gating properties of acetylcholine receptors (AChRs) change during the development of rat soleus endplates. During the first 3 weeks after birth, the apparent mean channel open time (tau) decreases severalfold and the single-channel conductance (gamma) increases 50%. To better understand this phenomenon, we used a combination of noise analysis, analysis of miniature endplate currents (MEPCs), and single-channel recordings to quantify the relative levels of fast and slow AChR activity at developing soleus endplates. When the same endplates were studied with both noise analysis and MEPC analysis, results obtained with the two techniques were strongly correlated, but MEPC analysis yielded higher estimates of the relative amount of slow channel activity. Experiments designed to examine the distribution of fast and slow channels gave no evidence for a gradient of either channel type within individual endplates; rather, fast and slow channels appeared to be mixed together. However, the relative amount of fast and slow channel activity did vary markedly among different endplates within individual muscles. In agreement with earlier studies, we found a progressive decrease in the relative amount of slow channel activity during the first 3 weeks after birth. However, our data indicate that this process begins sooner than reported previously and takes longer to complete. Some of the same endplates that were studied physiologically were also examined in the electron microscope to test the hypothesis that changes in AChR gating might be related to ultrastructural changes such as the formation of folds. The physiological and ultrastructural results were essentially uncorrelated.

2. At random sites on uninnervated, embryonic chick muscle fibres in vitro, T was relatively long -4 msec at 23 'C.3. Estimates of T at synaptic sites on embryonic myotubes innervated in vitro were identical to estimates at extrasynaptic sites on the same fibres. Both were comparable to estimates on uninnervated myotubes.

Incubating skeletal muscle fibers with forskolin, an activator of adenylate cyclase, increases the rate at which nicotinic acetylcholine receptors (AChRs) desensitize when exposed to ACh. Several reports indicate that this is due to the phosphorylation of AChRs by cAMP-dependent protein kinase, but other studies suggest that forskolin interacts with AChRs directly and that second-messenger systems are not required. To help clarify this issue, we studied the effects of forskolin and several other drugs on AChR function in embryonic rat myotubes. AChR function was studied by recording ACh-induced membrane depolarizations and ACh-induced single-channel currents. Our results indicate that forskolin at low concentrations enhances AChR desensitization through the action of a second messenger, most likely cAMP. An analog of forskolin that is much less effective in activating adenylate cyclase (1,9-dideoxyforskolin) is also much less potent in enhancing desensitization. Forskolin at low concentrations does not alter single-channel conductance or mean channel open time. However, when used at concentrations above 20 microM, forskolin may also exert direct drug effects on AChRs.

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