Distributions of active spinal cord neurons during swimming and scratching motor patterns

The spinal cord can generate motor patterns underlying several kinds of limb movements. Many spinal interneurons are multifunctional, contributing to multiple limb movements, but others are specialized. It is unclear whether anatomical distributions of activated neurons differ for different limb movements. We examined distributions of activated neurons for locomotion and scratching using an activity-dependent dye. Adult turtles were stimulated to generate repeatedly forward swimming, rostral scratching, pocket scratching, or caudal scratching motor patterns, while sulforhodamine 101 was applied to the spinal cord. Sulforhodamine-labeled neurons were widely distributed rostrocaudally, dorsoventrally, and mediolaterally after each motor pattern, concentrated bilaterally in the deep dorsal horn, the lateral intermediate zone, and the dorsal to middle ventral horn. Labeled neurons were common in all hindlimb enlargement segments and the pre-enlargement segment following swimming and scratching, but a significantly higher percentage were in the rostral segments following swimming than rostral scratching. These findings suggest that largely the same spinal regions are activated during swimming and scratching, but there are some differences that may indicate locations of behaviorally specialized neurons. Finally, the substantial inter-animal variability following a single kind of motor pattern may indicate that essentially the same motor output is generated by anatomically variable networks.     

Specification of synaptic connections between sensory and motor neurons in the developing spinal cord

Experimental studies of mechanisms underlying the specification of synaptic connections in the monosynaptic stretch reflex of frogs and chicks are described. Sensory neurons innervating the triceps brachii muscles of bullfrogs are born throughout the period of sensory neurogenesis and do not appear to be related clonally. Instead, the peripheral targets of these sensory neurons play a major role in determining their central connections with motoneurons. Developing thoracic sensory neurons made to project to novel targets in the forelimb project into the brachial spinal cord, which they normally never do. Moreover, these foreign sensory neurons make monosynaptic excitatory connections with the now functionally appropriate brachial motoneurons. Normal patterns of neuronal activity are not necessary for the formation of specific central connections. Neuromuscular blockade of developing chick embryos with curare during the period of synaptogenesis still results in the formation of correct sensory-motor connections. Competitive interactions among the afferent fibers also do not seem to be important in this process. When the number of sensory neurons projecting to the forelimb is drastically reduced during development, each afferent still makes central connections of the same strength and specificity as normal. These results are discussed with reference to the development of retinal ganglion cells and their projections to the brain. Although many aspects of the two systems are similar, patterned neural activity appears to play a much more important role in the development of the visual pathway than in the spinal reflex pathway described here.     

A change in the ultrastructure of frog spinal cord and motor neuron synapses during prolonged activation]

Motoneuron axosomatic synapses (AS) were shown to be larger than axodendritic (AD) synapses under anaesthesia. A 10 minute-long electrical stimulation both by low and high frequency caused enlargement of AS and AD synapses, it being the more pronounced the higher was the activation frequency. In all cases changes the AS synapses were more marked than those of AD synapses. On the bases of morphological and physiological data it was concluded that the changes of the synapse dimentions did not affect the synaptic transmission.     

Extensive fusion of mitochondria in spinal cord motor neurons

The relative roles played by trafficking, fission and fusion in the dynamics of mitochondria in neurons have not been fully elucidated. In the present study, a slow widespread redistribution of mitochondria within cultured spinal cord motor neurons was observed as a result of extensive organelle fusion. Mitochondria were labeled with a photoconvertible fluorescent protein (mitoKaede) that is red-shifted following brief irradiation with blue light. The behavior of these selectively labeled mitochondria was followed by live fluorescence imaging. Marking mitochondria within the cell soma revealed a complete mixing, within 18 hours, of these organelles with mitochondria coming from the surrounding neurites. Fusion of juxtaposed mitochondria was directly observed in neuritic processes at least 200 microns from the cell body. Within 24 hours, photoconverted mitoKaede was dispersed to all of the mitochondria in the portion of neurite under observation. When time lapse imaging over minutes was combined with long-term observation of marked mitochondria, moving organelles that traversed the field of view did not initially contain photoconverted protein, but after several hours organelles in motion contained both fluorescent proteins, coincident with widespread fusion of all of the mitochondria within the length of neurite under observation. These observations suggest that there is a widespread exchange of mitochondrial components throughout a neuron as a result of organelle fusion.     

The interaction between glycine and GABA postsynaptic effects in the spinal cord neurons of frog Rana temporaria

Patch-clamp study in the whole multipolar cell (presumably motoneuron) was performed, the cells having been mechanically isolated from the spinal cord of the frog Rana ridibunda. It was shown that GABA and glycine, when applied simultaneously, produced a transmembrane current. Its amplitude was lower than the summed amplitude of currents produced in the same neuron by separate applications of GABA and glycine. The investigation of this occlusion showed that the superfusion of the neurons with solution containing 0.2 mM of glycine totally blocked the responses to GABA (5 mM) application, and vice versa. The crossinhibition can lie in the basis of this phenomenon. It could be due to either the existence of a common receptor complex sensitive to both GABA and glycine or to interaction between GABA and glycine receptors.     

Relative numbers of neurons with different mediator mechanisms in motor nuclei of the cat cervical spinal cord

Motor nuclei of the cat cervical spinal cord are formed by groups of neurons differing in their mediator metabolism. From 40 to 65% are true motor (cholinergic) neurons. The localization of the precipitate of the reaction for acetylcholinesterase in the perinuclear space, on the membranes of the granular reticulum, axolemma, neurofilaments, and neurotubules of the axons, and in the synaptosomes and synaptic space are evidence of the possible perinuclear synthesis of the enzyme and of its transport with the flow of axoplasm. Comparison with the autoradiographic detection of glycine showed that large motor neurons form groups with small short-axon glycine-containing neurons, which make contact with them. The motor neurons have polyreceptive properties, for endings containing cholinesterase, glycine, noradrenalin, and serotonin, as well as unidentified endings are present on their soma and processes.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukranian SSR, Kiev. Translated from Neirofiziologiya, Vol. 9, No. 2, pp. 191–197, March–April, 1977.     

Chromatin organization in frog Purkinje neurons during the annual cycle: cytochemical and ultrastructural studies

Feulgen-DNA microdensitometric and propidium Iodide-DNA (PI-DNA) microfluorometric evaluations were made in Purkinje cells of cerebella from hibernating frogs and from active frogs. Higher Feulgen-DNA/distribution area ratios and lower PI-DNA values were found in the hibernating frog (both at high and low fluorochrome concentrations). These data indicate a higher degree of chromatin compactness. During the activity period, the Feulgen-DNA/distribution area ratios are lower and the Feulgen-DNA content is higher; the corresponding higher PI-DNA values indicate a lower degree of chromatin condensation. Histochemical and ultrastructural data show clearly different distribution patterns of chromatin and ribosomes in the two periods. The overall results underscore lower activity of neurons and a greater homogeneity within the Purkinje cell population during hibernation.     

Zebrafish transgenic constructs label specific neurons in Xenopus laevis spinal cord and identify frog V0v spinal neurons

A correctly functioning spinal cord is crucial for locomotion and communication between body and brain but there are fundamental gaps in our knowledge of how spinal neuronal circuitry is established and functions. To understand the genetic program that regulates specification and functions of this circuitry, we need to connect neuronal molecular phenotypes with physiological analyses. Studies using Xenopus laevis tadpoles have increased our understanding of spinal cord neuronal physiology and function, particularly in locomotor circuitry. However, the X. laevis tetraploid genome and long generation time make it difficult to investigate how neurons are specified. The opacity of X. laevis embryos also makes it hard to connect functional classes of neurons and the genes that they express. We demonstrate here that Tol2 transgenic constructs using zebrafish enhancers that drive expression in specific zebrafish spinal neurons label equivalent neurons in X. laevis and that the incorporation of a Gal4:UAS amplification cassette enables cells to be observed in live X. laevis tadpoles. This technique should enable the molecular phenotypes, morphologies and physiologies of distinct X. laevis spinal neurons to be examined together in vivo . We have used an islet1 enhancer to label Rohon‐Beard sensory neurons and evx enhancers to identify V0v neurons, for the first time, in X. laevis spinal cord. Our work demonstrates the homology of spinal cord circuitry in zebrafish and X. laevis , suggesting that future work could combine their relative strengths to elucidate a more complete picture of how vertebrate spinal cord neurons are specified, and function to generate behavior. © 2017 Wiley Periodicals, Inc. Develop Neurobiol 77: 1007–1020, 2017     

Possible correlations between the level of alcoholic motivation and the electrophysiological structure of sleep in the rat

Using modified Porsolt's method, the electrophysiological sleep pattern was studied in normal conditions and after a single intraperitoneal ethanol injection to noninbred male albino rats divided into 2 groups ("high activity" and "low activity" rats). Voluntary alcohol intake in these rats was measured during free choice between 10% ethanol and water for 20 days. "Low activity" rats were characterized by a statistically significant 3.4-fold higher level of ethanol consumption and 2.7-fold longer REM-sleep stage, as compared to "high activity" animals. In "low activity" animals ethanol (1 g/k, 10% solution, i. p.) inhibits and in "high activity" rats it increases REM-sleep stage, thus removing differences in the sleep pattern in the two groups of rats. The data obtained suggest a possible role of REM-sleep in the development of alcohol motivation.     

Electrotonic transmission between primary afferent fibers and the motor neurons of the isolated spinal cord of various representative amphibians

Studies have been made of monosynaptic excitatory postsynaptic potentials elicited by stimulation of the posterior root in motoneurones of the isolated spinal cord of the frog Rana ridibunda, toad Bufo bufo, and clawed toad Xenopus laevis. In all the amphibians studied, the early component of monosynaptic EPSP was not blocked in Ca-free medium containing 2 mM Mn2+. It is suggested that electrical coupling in anuran amphibians reflects certain stage of evolution of the synaptic transmission in vertebrates.     

Morphologic and cytophotometric indices of the state of the neurons of the spinal cord and spinal cord ganglia following exposure to gravitational stress]

The work presents data on the state of the neurons of the spinal ganglia and the spinal cord of the lumbosacral part in 96 cats subjected to single stresses (10g) and repeated ones (6g). The material was stained with thionin after Nissl. RNA was detected after Einarson. Photometry of the sections was made in MUP.5. Morphological changes were shown to be more pronounced in sensory cells. The method of cytophotometry established the increase by 25% amount of RNA in their cytoplasm after single stresses. Repeated stresses resulted in depletion of RNA. The amount of RNA returned to the initial level within 2 days after single stresses and within 3 days after repeated rotations. The shifts were much less pronounced in motoneurons.     

Synaptic interaction between single motoneurons in the isolated frog spinal cord

The nature of synaptic interaction between two neighboring motoneurons in the isolated frog spinal cord was studied by parallel insertion of two separate micro-electrodes into the cells. In 82 of 89 motoneurons tested transmission through synapses between the motoneurons was electrical in nature, as shown by the absence or short duration of the latent period of elementary intermotoneuronal EPSPs, stability of their amplitude, and preservation of responses in Ca⁺⁺-free solution containing 2 mM Mn⁺⁺. Direct electrotonic interaction was demonstrated in both directions: artificial de- and hyperpolarization of one motoneuron led to corresponding shifts of membrane potential in the neighboring motoneuron. The time constant of rise and decay of this potential was appreciably greater than the time constant of the membrane of the two interconnected motoneurons. Blockade of the SD-component of the action potential in the "triggering" motoneuron led to a decrease in the elementary EPSP in the neighboring motoneuron. These facts suggest that electrotonic interaction takes place through dendro-dendritic junctions. Absence of rectification was demonstrated in electrical synapses between motoneurons. In four cases elementary EPSPs were chemical in nature, for they appeared 1.3–3.3 msec after the beginning of the action potential in the "triggering" motoneuron, and were blocked in Ca⁺⁺-free solution containing Mn⁺⁺; fluctuations of their amplitude approximated closely to a Poisson or binomial distribution. Such responses are evidently generated by synapses formed by recurrent axon collaterals of one motoneuron on the neighboring motoneurons. In three cases elementary intermotoneuronal EPSPs consisted of two components, the first electrical and the second chemical in nature. Morphological structures which may be responsible for generation of 2-component EPSPs are examined.Deceased.I. M. Sechenov Institute of Evolutionary Physiology and Biochemistry, Academy of Sciences of the USSR, Leningrad. Translated from Neirofiziologiya, Vol. 16, No. 5, pp. 619–630, September–October, 1984.