In vivo development of nicotinic acetylcholine receptor channels in Xenopus myotomal muscle

Abstract

The development of acetylcholine receptor (AChR) channel function in Xenopusmyotomal muscle was studied by singlechannel recordings from cell-attached patches of nonjunctional membrane in viva. AChR channels were studied from the time of their first appearance on the muscle membrane until the time of full maturity of the muscle. Two predominant amplitude classes of AChR channels were observed with slope conductances of about 40 and 60 pS. During the first day after their initial appearance on the membrane, the smallconductance channels were the most numerous class on the muscle membrane.The large-conductance channels then began to be expressed in significant numbers and, over the next 2 d, became the predominant channel type. The largeconductance class had an apparent mean open time of -0.7 msec at resting potential, which remained constant throughout development.The small channel initially had an apparent open time of -3 msec at resting potential, which decreased during development by about 50%. The decrease in open time of the small channel was correlated in time with the increased expression of the large-conductance channels. Openings of the large-conductance channels were generally separated by closed intervals of more than 1 msec, whereas openings of the small-conductance channels were commonly interrupted by brief gaps of about 0.2 msec duration. The duration of the brief gaps did not change during development.Studies of amphibian and mammalian skeletal muscle have shown that nicotinic acetylcholine receptor (AChR) channels undergo functional changes during development (reviewed by Schuetze and Role, 1987). In general, there is an increase in AChR channel conductance and a decrease in open time as the muscle matures. The first evidence of such changes came from the analysis of ACh-induced noise recorded from muscle. Recordings from muscle membrane in rats (Sakmann and Brenner, 1978;Fischbach and Schuetze, 1980) and amphibia (Kullberg et al., 198 1;Kullberg and Kasprzak, 1985) demonstrated a 3-to 4-fold decrease in the open time of AChR channels. These experiments revealed the chronology of change in gating during muscle development, but they gave no information about the conductances of the channels. With the advent of the single-

Multiple conductance classes of mouse nicotinic acetylcholine receptors expressed in Xenopus oocytes.

“…Recordings from BC3H-1 cells (27) have not revealed multiple conductance levels of AcChoRs such as we observed in oocytes; however, AcChoR channels with conductances lower than 40 pS have been reported in Xenopus muscle in vivo (7,8,28) and in denervated mouse muscle (29). One of these, a 25-pS channel, has been well characterized in developing Xenopus muscle (8).…”

“…Single channel events were recorded by a List EPC/7 patch clamp amplifier, and stored on FM tape for off-line analysis. The procedures for data analysis were similar to those described (7). Records from a8y3-injected oocytes were low-pass Bessel …”

“…Although both kinds of receptors assemble less efficiently and the 8-less receptors have reduced agonist affinity (5), the weak responses to AcCho might also reflect a reduced conductance or a briefer open time. We were interested in examining the properties ofthese receptors because they might offer an explanation for the small conductance channels (10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25) pS) observed in developing amphibian muscle (7,8). We injected mRNAs encoding the mouse a, 3, y, and 8 subunits into Xenopus oocytes and found that multiple conductance classes of channels were expressed on the oocyte membrane, and one of these, the smallest, is due to deletion of the 8 subunit.…”

Potentiation by ATP of the postsynaptic acetylcholine response at developing neuromuscular synapses in Xenopus cell cultures.

“…However, at the single-channel level, our results showed that ATP had a selective action on the low-conductance ACh channel, a characteristic only of embryonic muscle cells. Developmental changes of ACh receptor channels have been studied extensively in Xenopus muscle cells in vitro as well as in vivo (Brehm, Kidokoro & Moody-Corbett, 1984;Owens & Kullberg, 1989;Rohrbough & Kidokoro, 1990). Shortly after the initial insertion of ACh receptor channels a great majority of the channels are of an embryonic type (lowconductance channels), which have a prolonged mean open time and low conductance.…”

Acetylcholine receptor channels in Xenopus myocyte culture; brief openings, brief closures and slow desensitization.

“…Developmental changes of acetylcholine (ACh) receptor channels have been studied extensively in Xenopus myotomal muscle cells in vitro (Kidokoro, Brehm & Gruener, 1982; Brehm, Kidokoro & Moody-Corbett, 1984 a;Igusa & Kidokoro, 1987; Leonard, Nakajima, Nakajima & Carlson, 1988) as well as in vivo (Kullberg, Brehm & Steinbach, 1981;Brehm, Kullberg & Moody-Corbett, 1984b; Owens & Kullberg, 1989). Shortly after initial insertion of ACh receptor channels a great majority of channels are an embryonic type which have a prolonged mean burst duration.…”