1. The inferior rectus muscle of rat, one of the extraocular muscles, contains two populations of multiply innervated fibers (MIFs): orbital MIFs, located in the orbital layer of the muscle and global MIFs, found in the global layer. The electrical properties and the responses to nerve stimulation of orbital MIFs were studied with single intracellular electrodes and compared with those of twitch fibers of the orbital layer, MIFs of the global layer, and tonic fibers of the frog. 2. About 90% of the orbital MIFs did not produce overshooting action potentials. In these fibers the characteristics and time course of the responses to nerve stimulation varied along the length of the fibers. Within 2 mm of the end-plate band of the muscle, the responses consisted of several small end-plate potentials (EPPs) and a nonovershooting spike. Distal to 2 mm, the responses in most fibers consisted of large and small EPPs with no spiking response. Some fibers produced very small spikes surmounted on large EPPs. 3. Overshooting action potentials were observed in approximately 10% of the orbital MIFs recorded between the end-plate band and 2 mm distal. The presence or absence of action potentials was not related to the magnitude of the resting potential of the fibers. 4. The threshold of nerve stimulated responses in orbital MIFs was the same as that in orbital twitch fibers. A large number of orbital MIFs had latencies equal to those for the orbital twitch fibers recorded at the same distance from the end-plate band, but the average latency was greater in the MIFs. The latency of orbital MIFs was about one-half of that for the MIFs of the global layer. The values for the effective resistance and membrane time constant of orbital MIFs fell between those for orbital twitch fibers on the one hand, and global MIFs and frog tonic fibers on the other. 5. In order to compare electrical properties with innervation patterns, fibers identified electrophysiologically as orbital MIFs were injected with the fluorescent dye Lucifer yellow and then traced in Epon-embedded, serial transverse sections. In addition to numerous superficial endings distributed along the fibers, a single "en plaque" ending was also found in the end-plate band that resembled the end plates of the adjacent orbital twitch fibers. 6. From these results we conclude that the electrical activity of orbital MIFs varies along the length of the fibers.(ABSTRACT TRUNCATED AT 400 WORDS)

Monoclonal antibodies (McAB) specific for fast (C14) and slow (S58) myosin, and a myosin antigenically similar to neonatal/embryonic myosin in mammals (ALD180), were used to characterize the myosin distribution in orbital layer fibres of rat extraocular muscles (EOM) in relation to innervation patterns. The orbital layer is composed of both singly-innervated (SIF) and multiply-innervated (MIF) fibres. The SIFs have the characteristics of twitch fibres, while the MIFs, in addition to possessing many small endings characteristic of tonic fibres, also have an en-plaque-like innervation in the endplate band resembling that of the adjacent SIFs. Myosin expression in MIFs and SIFs is unusual and varies systematically along the length of the fibres. Both SIFs and MIFs label with ALD180, but this labelling is absent in both fibre types in the endplate band region, where all fibres label with C14. Distally and also proximally to the endplate band, SIFs label with both ALD180 and C14, while the MIFs, innervated by many small, superficial endings in these regions, label with ALD180 only. This pattern of myosin expression could also be demonstrated in isolated fibres. The results are discussed in relation to the hypothesis that both populations of orbital layer fibres express constitutively both fast and the neonatal-like myosin, and that superimposed on this constitutive expression twitch or tonic innervation acts locally to selectively suppress either neonatal-like or fast myosin, respectively.

Extraocular muscles contain both fast-twitch and multiply-innervated, tonic-contracting fibres. In rat, these fibres collectively express numerous myosin heavy chain isoforms including fast-type embryonic and neonatal, adult slow twitch type I and fast twitch type II, and a fast isoform unique to extraocular muscle. Immunocytochemical and Western blotting results are presented which suggest that, in rabbit, an additional species, the alpha-cardiac myosin heavy chain, is present. The immunoreactive species is found in all rabbit extraocular muscles and in the extraocular muscles is expressed in almost all fibres which do not contain a fast myosin heavy chain. Positive identification of this isoform as the alpha-cardiac myosin heavy chain was obtained by sequencing a cloned PCR product derived from extraocular muscle mRNA unique to the 3'-end of rabbit alpha-cardiac myosin heavy chain mRNA. This is the first unequivocal demonstration of alpha-cardiac myosin heavy chain expression in extraocular muscle.

The transition from aquatic to terrestrial hearing in the frog occurs during metamorphosis and during the disappearance of the lateral line system. The coincidence in time of these two processes and morphological similarities between the acoustic and lateral line systems has led to the suggestion (Larsell, '34) that the lateral line nuclei are transformed into the acoustic nuclei. The relation between the acoustic and lateral line systems was investigated by studying the distribution of primary afferents, the dendritic patterns of the cells in the primary nuclei, and the development of the nuclei in the premetamorphic bullfrog, Rana catesbeiana. The posterior and anterior lateral line roots distribute to a neuropil located medial to the dorsal medullary nucleus. Horseradish peroxidase (HRP) injections into the contralateral tegmentum fill cells in the periventricular region whose dendrites ramify within the neuropil. These cells constitute the lateral line nuclei. The amphibian and basilar papillary roots of the acoustic system distribute to the more lateral nuclear region. The dendrites of these cells arborize within the nucleus and not in the lateral line neuropil. The dorsal medullary nucleus is, therefore, the acoustic nucleus (AcN). [3H]-thymidine labeling reveals that newly generated cells occupy the AcN within a few hours of their formation throughout the period when anatomical analysis shows the parallel growth and diminution of the lateral line neuropil and nuclei. This study indicates that the lateral line and acoustic systems are morphologically independent at the level of the primary afferents and primary nuclei throughout early development.

The properties of extraocular muscles (EOMs) are quite different from those of the trunk and limb. Here we show that there is a novel pattern of troponin T (TnT) expression in EOMs which most likely contributes to the fine control of ocular movement and may reflect their innervation by cranial motoneurons. Three regions of the muscle were analysed to distinguish the TnT isoforms present in the fast singly-innervated fibres from those in the multiply-innervated fibres. More than 95% of the TnT in the singly-innervated fibres is TnT3f, which exhibits the most graded response to changes in calcium concentration during activation (Schachat et al., J. molec. Biol. 198, 551-4). In multiply-innervated fibres, which exhibit tonic contractures, the slow troponin T TnT2s is expressed. While neither TnT3f nor TnT2s is unique to EOM, this pattern is unusual in two respects: first, both TnT3f and TnT2s are minor components of the trunk and limb musculature, and second, most muscles express several fast and both slow TnT species. Although EOM occupies a highly specialized physiological niche, its unusual physiology is not reflected in the presence of new TnT isoforms but in the expression of a different ratio of the known species of TnT.

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We describe the relation between growth and branching of an identified dendrite and the formation of synapses on its surface during a 3 1/2-day period early in development. We studied the lateral dendrite and the adjacent lateral perikaryon of the Mauthner cell (M-cell) during embryonic stages 39-43 in the axolotl Ambystoma mexicanum. Reconstructions from light micrographs of serial sections through the cell revealed that during this interval the dendrite elongates rapidly, and large numbers of ventrally directed branches are formed. Samples of the same material by electron microscopy showed that large numbers of synaptic contacts appear during the same interval. We quantitatively estimated changes in local synapse densities (the number of contacts/100 micrometers2 of M-cell surface) and local surface areas of the M-cell and found that synapses were most densely clustered, and accumulated most rapidly, on regions of the cell that were rapidly expanding. These data are in accord with previous evidence from work in this and in other systems that synaptic contacts induce local growth of dendrites. Furthermore, the data are consistent with a proposal that outgrowth of new dendritic branches is induced or stabilized by synapses in a concentration-dependent fashion.

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