Organization and function of molecular domains in myelinated neurons

Myelinated neurons are organized into distinct molecular domains that are essential for electrical conduction. These domains include the axon initial segment, nodes of Ranvier, paranodes, juxtaparanodes, and internode. The paranode acts as a molecular fence, maintaining separation of the sodium channel-enriched nodes from the potassium channel-enriched juxtaparanodes via axo-glial septate junctions. In demyelinating diseases like multiple sclerosis and Guillain-Barré, the clustering and segregation of such domains and structures are disrupted, resulting in altered neuronal function. Whirlin (Whrn) is a PDZ-containing cytoskeletal protein whose role is to cluster and segregate transmembrane proteins in both the ear and eye. We examined Whrn knockout (Whrn-/-) mice and how the loss of Whirlin protein affected myelinated axonal domain organization. Our studies confirmed loss of the major 110kDa Whrn isoform in Whrn-/- dorsal root ganglia, spinal cord, and eye tissue lysates. Curiously, the expression levels of nodal, paranodal, and juxtaparanodal markers in spinal cord lysates were similar between wild-type and Whrn-/- mice. Also, conduction velocities in isolated Whrn-/- sciatic nerves were similar to wild-type. We detected and quantified a `spring-like' paranodal compaction phenotype in four to eight week-old Whrn-/- sciatic nerves. The paranodal compaction phenotype, juxtaparanodal and associated cytoskeletal disruption were observed in Whrn-/- mutant sciatic nerves and spinal cord fibers from two week-old to one year-old mice. Light and transmission electron microscopic analyses of Whrn-/- mice revealed cellular swellings in cerebellar Purkinje axons filled with mitochondria and vesicles. Such Purkinje axon findings were similar to the Caspr, another paranodal protein, knockout mouse cerebellar phenotype. Additional higher magnification, electron micrograph images of the paranodal, juxtaparanodal and internodal region in Whrn-/- mice reveal abnormal accumulation of mitochondria and vesicles. Taken together, our studies revealed Whirlin plays an important role in paranodal and juxtaparanodal organization, maintaining cytoskeletal organization, and preventing accumulation of mitochondria and vesicles. In addition to these core findings, results are included from a study characterizing fluorescent imaging of myelination/demyelination in the cuprizone mouse model, as well an investigation of the localization and function of PDZD7 in the inner ear, a protein structurally similar to Whirlin.Doctor of Philosoph