Connexins in Neuronal and Glial Gap Junctions in the Central Nervous System

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DESCRIPTION (provided by applicant): Gap junctions are microscopic intercellular junctions that provide for direct intercellular ionic, electrical, and metabolic coupling between nerve cells in the brain, retina, and spinal cord. Previously, gap junctions between neurons were thought to consist of hundreds or thousands of connecting channels ("connexons"), but they were also thought to be rare, to occur only between a few types of neurons, and to occur only in limited, non- cognitive areas of the central nervous system. The discovery of "miniature" gap junctions (<100 connexons) and preliminary evidence for their abundance throughout the brain suggests that "mini" gap junctions, particularly those at "mixed" (chemical plus electrical) synapses, may provide the structural basis for the detection of tiny sub-threshold electrical "spikelets" or "partial spikes" in principal neurons that are distributed throughout the brain. Gap junctions / partial spikes are thought to be essential for regulating and optimizing the high-frequency neuronal oscillatory synchronizations that are thought to underlie consciousness, arousal from sleep, cognition, associative binding for learning and memory, and fine motor control, and which become pathologically altered in epilepsy, schizophrenia, Parkinson's disease, and autism spectral disorders. We will combine laser scanning confocal immunofluorescence microscopy with newly-developed freeze-fracture replica immunogold labeling (FRIL) electron microscopy to detect, quantify, determine the protein composition of, and measure the sizes of gap junctions throughout the complex circuitry of the mouse brain, to make detailed measurements of "mini" gap junctions in cerebral cortex and hippocampus of both mouse and human brain, and to identify the neuronal subtypes linked by "mini" gap junctions. We will emphasize analysis of those regions that are primarily responsible for thinking and consciousness (cerebral cortex) and for learning and memory (hippocampus) and that represent the primary sites of origin of epileptic discharges. These complementary approaches will allow direct correlation of data from large-scale (whole mouse brain) to ultrastructural and molecular levels. The data to be obtained will be essential for understanding how consciousness is created, what distinguishes sleeping vs. awake states, how consciousness is altered during general anesthesia, and how memories are created, and will also be essential for identifying subcellular sites that may become targets for designing new drugs to treat disorders of electrical synaptic communication in the brain. PUBLIC HEALTH RELEVANCE A newly-discovered class of "miniature" gap junctions, found to be abundant between many classes of neurons, may provide the structural basis for propagation of small electrical "spikelets" between neurons, and thereby, to regulate the oscillatory synchronizations of cerebral cortex and hippocampus that are associated with human consciousness and for associative binding for learning and memory. We will detect, map, quantify, and determine the connexin protein composition of "mini" gap junctions throughout mouse brain, both at electrical and at "mixed" (chemical plus electrical) synapses, and compare the distributions of gap junctions and their constituent connexins proteins in mouse vs. normal human cerebral cortex and hippocampus. These data will serve as a prelude to measuring changes in these structures in human disease such as epilepsy, schizophrenia, Parkinson's disease, and autism spectral disorders.