Correlated muscle and nerve development in the bullfrog cutaneous pectoris.

The development of the cutaneous pectoris muscle was studied and compared with the differentiation of its peripheral nerve in bullfrog (Rana catesbeiana) tadpoles and frogs by light and electron microscopic techniques. This muscle preparation was chosen for this study because it possesses a number of advantages for (and has become a model system for) the study of correlated nerve-muscle development. At the earliest stage examined (stage XI) the presumptive muscle did not contain any contractile or morphologically distinguishable myotubes, but was contacted by the well-defined cutaneous pectoris nerve trunk. Myotubes were present at stage XII, the same time that nerve-associated acetylcholine receptor aggregations and nerve-evoked muscle contractions were first observed. The adult number of axons was present in the cutaneous pectoris nerve at stage XII, but no axons were myelinated. Gradually thereafter, the number of muscle fibers increased and the cutaneous pectoris axons became myelinated. By stages XX and XXI, but prior to metamorphic climax (stage XXV), the adult numbers of cutaneous pectoris muscle fibers and myelinated and unmyelinated nerve fibers were present. These numbers did not change significantly between stages XX and XXI, through metamorphosis, and in the adult, even during the period of the most rapid loss of multiple innervation in the first 2 weeks after metamorphosis. These results show that the nerve was present and in contact with the cutaneous pectoris muscle from the earliest stages of development prior to muscle differentiation, at a time when the muscle was a disorganized mass of undifferentiated cells. Such early contact suggests that the nerve may have a significant influence on muscle maturation.

Neuromuscular junction development in the cutaneous pectoris muscle of Rana catesbeiana.

Synaptic specializations were studied in the developing cutaneous pectoris muscle of Rana catesbeiana tadpoles and froglets to correlate nerve terminal morphology (by light and electron microscopy), accumulation of acetylcholine receptors, and the ability of the muscle to contract following nerve stimulation. This correlated approach was used to determine the developmental timing and possible causal relationship of events in nerve and muscle maturation at the neuromuscular junction. Initially, the cutaneous pectoris nerve trunk was present in the undifferentiated presumptive cutaneous pectoris mesenchyme, prior to muscle maturation. At stage XII when the muscle was first able to contract weakly in response to nerve stimulation, the motor nerve terminal endings were simple bulbous enlargements associated with diffuse subneural aggregations of acetylcholine receptors (indicated by diffuse speckles of rhodamine alpha-bungarotoxin fluorescence). Before stage XII no rhodamine alpha-bungarotoxin fluorescence was present anywhere in the muscle. The first stage in the organization of acetylcholine receptors at the neuromuscular junction was the accumulation of diffuse speckles of fluorescence beneath the terminal enlargements. This was followed by the clustering of receptors into small polygonal areas at each synaptic site, and finally the organization of receptors into parallel linear rows. Presumably this final stage was associated with formation of junctional folds. By stage XV the synapses were multiply innervated and had developed acetylcholinesterase activity. The general nerve terminal morphology and pattern of accumulation of acetylcholine receptors at cutaneous pectoris neuromuscular junctions were similar to those of the adult throughout metamorphic climax except that they still contained more than one motor axon. After metamorphic climax, elimination of multiple innervation occurred.

Rapid decline in acetylcholine release and content of rat extensor digitorum longus muscle after denervation.

The amount of acetylcholine (ACh) and choline (Ch) in normal and denervated rat extensor digitorum longus (EDL) muscles, as well as that released spontaneously from these muscles, was determined by an extremely sensitive gas chromatographic-mass spectrometric assay method. We found decreases in ACh content and spontaneous, resting ACh release as early as 8 h after denervation. The ACh content decreased to a plateau of 30% of control by 11 h; ACh release attained a plateau of 50% of control several hours later. These results showed that in denervated EDL muscles ACh content and spontaneous release (measured biochemically) decreased before nerve-evoked and spontaneous quantal release (measured physiologically) ceased at most synapses. The rapid reduction in ACh, or possibly in other substances that may be released with ACh, may be an important factor in initiating postsynaptic degenerative changes after nerve transection. Choline content and choline resting release increased significantly at both 1 and 3 days after nerve transection. These increases may be related to onset of postsynaptic neuromuscular degenerative changes.

The dorsal lateral geniculate nucleus of the normal ferret and its postnatal development.

The anterograde transport of 3H proline and of horseradish peroxidase has been used to study the retinogeniculate pathway in normal adult ferrets and in young ferrets during postnatal development. the lateral geniculate nucleus in adults shows a characteristic "carnivore" pattern, with layers A, A1, C, C1, C2, and C3, and a medial interlaminar nucleus recognizable either cytoarchitectonically or on the basis ofth retinogeniculate innervation. In addition, there is a well-defined, rather large perigeniculate nucleus. At birth the lateral geniculate nucleus is unlaminated and essentially all parts are reached by afferents from both eyes. The crossed component is by far the larger. It extends from the optic tract medially well into the perigeniculate field, in contrast to the uncrossed component which barely reaches the perigeniculate field. During the first 3 postnatal days the uncrossed fibers restrict their arbors to a small posterior and medial region, the precursor of the biocular segment of the nucleus. The crossed fibers gradually retreat from the region within which the uncrossed fibers have concentrated. Between the fourth and eighth postnatal days the field occupied by the ipsilateral component expands again to form a major focus that will define lamina A1 and a minor focus that will define C1. At this stage the crossed and the uncrossed fibers overlap at the borders of lamina A1 and the whole region of lamina C1 is also occupied by arbors of the crossed component. The perigeniculate field becomes clearly distinguishable from the lateral geniculate nucleus and the medial interlaminar nucleus is becoming clearly recognizable between days 3 and 8. Between days 8 and 15 the cytoarchitectonic borders between layers A and A1 become clearly defined, but the retinogeniculate axons from each eye still extend across this border. These axons retreat into their appropriate lamina after the 15th postnatal day an the nucleus reaches its essentially adult structure by about the fourth postnatal week. Segregation of retinofugal axons in the C layers occurs after segregation in the A layers, but many of the cells within the C layers show signs of cytological maturity earlier than those of the A layers. The nucleus undergoes a series of migrations and changes of shape as the ipsilateral and contralateral components become segregated. Whereas in teh newborn the nucleus is roughly comma-shaped and on the lateral aspect of the dorsal thalamus, in the adult it is "L"-shaped and mainly on the posterior aspect of the dorsal thalamus.