Our long term goal is to gain a better understanding of the functional organization of the mammalian spinal cord. Until recently, neurons in the adult mammalian spinal cord were thought to be linked only by chemical synapses. However, direct electrical communication via """"""""gap junctions"""""""" is known to occur between neurons in limited regions of the mammalian brain, and electrical coupling of spinal cord motor neurons has been detected in adult rats and cats. Nevertheless, extensive searches by conventional thin-section electron microscopy have failed to detect gap junctions between neurons in the spinal cords of adult mammals. (The only documented exception has been the demonstration of gap junctions in a small subset of testosterone-regulated neurons innervating sexually dimorphic muscles in the adult rat [i.e., the spinal nucleus of the bulbocavernosus muscle]).. In contrast, our newly-developed """"""""grid-mapped freeze-fracture"""""""" technique has revealed that numerous very small gap junctions occur on neuron somata, dendrites, and synaptic boutons throughout the ventral horn of the spinal cord in male and female rats. Because of the potential significance of gap junctions to normal spinal cord function, as well as to possible routes of intervention following spinal cord injury, our general aim is to determine the frequency and distribution of gap junctions among the various classes of motor neurons and interneurons in the spinal cord.
Our specific aims are: a) to utilize newly-developed """"""""grid-mapped freeze- fracture"""""""" to systematically map the occurrence and distribution of gap junctions within specific motor nuclei and intermediate gray interneurons in ventral and dorsal horns of the lumbosacral and cervical enlargements and thoracic cord segments of adult male and female rats, b) to determine the size, number, and subcellular distribution of those neuronal gap junctions, and c) to determine the functional identities of the spinal cord neurons that are linked by gap junctions. We will use laser confocal microscopy, """"""""grid-mapped"""""""" freeze-fracture, and """"""""sectioned-replica"""""""" electron microscopic techniques, combined with retrograde and anterograde tracing techniques and immunocytochemical labeling of gap junctions, to identify and map the subcellular distribution of gap junctions in identified subsets of alpha- and gamma-motor neurons and in the interneurons of discrete regions of the spinal cord (i.e., the ten """"""""laminae of Rexed""""""""). Identification and mapping of the neuronal cell types linked by gap junctions will confirm the existence of pathways for integration. and/or modulation of motor activity additional to those currently recognized. The data obtained in this research will be of value in a) understanding spinal cord synaptic organization, including a possible component of """"""""pattern generator"""""""" circuits, b) clarifying the functional organization of specific neuronal pools, and c) identifying- potential intercellular pathways for transfer of """"""""second messengers"""""""" in long-term trophic regulation of spinal cord neurons' in health, trauma, and disease.
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