Developmental changes in the distribution of acetylcholine receptors in the myotomes of Xenopus laevis.

Abstract

SUMMARY1. The acquisition and distribution of nerve fibres and of acetylcholine (ACh) receptors were examined in the myotomes of Xenopus laevis during normal development. This muscle is well-suited for investigating temporal relationships during neuromuscular synaptogenesis because the age of the Xenopus embryo at the onset of innervation can be assessed with an accuracy of about one hour. Myotomal nerve fibres were visualized after staining them with nitroblue tetrazolium and ACh receptors were examined after exposure to x-bungarotoxin labelled with 1251 or fluorescent dye.2. Nerve fibres were seen in the myotomes of some embryos as early as stage 19(20'75 hr) and in virtually all embryos by stage 24 (26-25 hr). From the outset they were located mainly at the ends of the myotomes, but some myotomes also exhibited nerve fibres in more central regions.3. ACh receptors were already present in myotomes by stage 19 (20-75 hr) and initially had a widespread, uniform distribution. The density of extrajunctional ACh receptors increased until stage 36 (50 hr) and then declined less than 3-fold over the next 10 days of development.4. Discrete patches of high ACh receptor density began to appear at the ends of the myotomes at stage 22 (24 hr) and were seen in almost all embryos by stage 26 (29-5 hr). ACh receptor patches were also seen in central regions of some myotomes and these were usually aligned in patterns which resembled the course of nerve fibres.5. The present findings suggest that myotomal muscle cells in Xenopus embryos begin to acquire ACh receptors shortly before the arrival of nerve fibres and that discrete patches of ACh receptors begin to form at presumptive synaptic sites on the average about 3 hr after the arrival of the nerve fibres. The latter delay is considerably shorter than that in developing rat muscle.6. The temporal and spatial relationships between nerve fibres and the development of ACh receptor patches in Xenopus myotomes in vivo are consistent with findings in Xenopus cell cultures which indicate that nerve fibres can rapidly induce ACh receptor localization at sites of nerve-muscle contact.

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

“…AChR single-channel activity was recorded from myotomal muscle in embryos as young as 2 1 hr (stage 19), the earliest age at which nicotinic AChRs are expressed on the muscle surface (Blackshaw and Warner, 1976;Kullberg et al, 1977;Chow and Cohen, 1983). Records were taken at each stage of development between stages 19-25 (a 6 hr period) and at about every third or fourth stage over the next 25 d (stages 25-54).…”

“…At very young stages (19)(20)(21)(22)(23)(24) or older stages (>48), some patches had no identifiable channel activity or else the event frequency was too low for analysis. This variability may reflect a lower density of nonjunctional AChR channels in very young and mature muscle relative to intermediate stages (Chow and Cohen, 1983). Single-channel conductances When multiple openings were eliminated, the single-channel currents fell into 1-4 different amplitude classes.…”

“…The cells of the myotomal muscle are short fibers, ranging in length from 100 pm at the time of synapse formation (stage 21) to about 300 pm at early metamorphosis (stage 48-50). The cells are arranged in myotomes along the length of the tail, and synapses are located oi each end of the cell-near the-myocommata (Kufibeig et al, 1977: Chow. 1980: Chow and Cohen.…”

“…The cells are arranged in myotomes along the length of the tail, and synapses are located oi each end of the cell-near the-myocommata (Kufibeig et al, 1977: Chow. 1980: Chow and Cohen. 1983.…”

Localization of cholinesterase at sites of high acetylcholine receptor density on embryonic amphibian muscle cells cultured without nerve

“…In skeletal muscle, acetylcholine receptors (AChRs) and cholinesterase (ChE) are both present in high density at the neuromuscular junction and become localized there very early during the development of the synapse (Diamond and Miledi, 1962;Bennett and Pettigrew, 1974;Bevan and Steinbach, 1977;Kullberg et al, 1977Kullberg et al, , 1980Chow, 1980). That nerve can induce the localization of both of these molecules at sites of innervation has been revealed in cultures of embryonic nerve and muscle cells Frank and Fischbach, 1979;Rubin et al, 1979) and in studies on the reinnervation of adult muscle in uivo (Aitken, 1950;Miledi, 1962;Fex et al, 1966;Lomo and Slater, 1980).…”