Our research uses the technique of extracellular patch clamping in the exploration of the role of ion channels in non-excitable cells. Ion channels are intrinsic membrane proteins that mediate fast communication in nerve and muscle. They are also important to a myriad of functions of non-excitable cells. Our goal is to develop a greater understanding of the relationship of ion channels, regulation of intracellular Ca2+ levels, and secretion in cells as diverse as phagocytic macrophages, inflammatory microglia, myelinating oligodendrocytes and neurons.

 Recent work has identified cDNAs encoding both delayed rectifier-type as well as inwardly rectifying K+ channels in human neurons. Expression studies in Xenopus oocytes and stable cell lines have confirmed the similarity of these currents to those observed in normal cells. CDNAs of the outwardly rectifying K+ channels encode subunits that can arrange to form active heteromultimers which may also be an important mechanism in generating the diversity of K+ currents present in a variety of tissues such as brain, heart, and pituitary. Antibodies directed against the channel and a fusion epitope have allowed characterization of the protein in transfected cell lines and transgenic animals. Additional studies have examined the mechanisms of subunit association and voltage-dependent inactivation in members of the delayed rectifier-type K+ channel family.

 Several members of the inwardly rectifying class of K+ channels have been cloned and expressed. Our laboratory has also identified several new cDNAs and their genes encoding new members of this family including the human ROMK1 ATP-regulated inward rectifier and GIRK1, a G-protein linked inward rectifier. Mutagenesis studies of GIRK1 have allowed us to identify the subunit assembly site on the C-terminal domain of the protein and structurally model the multi-protein interactions. Mutagenesis and expression studies of another prominent member of the same inward rectifier K+ channel family, IRK1, cloned from macrophages and also prominently expressed myelinating oligodendrocytes, have allowed us to determine those residues which are important in determination of the permeation properties of the pore of the channel. Protein modeling of this region will allow us to make predictions concerning the possible structure and assembly of multi-ion pores of which IRK1 is an example. These studies will extend to the molecular structures underlying the gating mechanisms of the channels and to the binding sites for external ligands (drugs) of possible clinical relevance. This may allow rational drug design to modify the function of ion channels in patients with inflammatory diseases such as multiple sclerosis, ischemic injury to the brain as well as other functions disordered by disease. The molecular mechanism of ion channel modulation by a diversity of stimuli including adrenergic mediators and cytokines can be resolved by the use of the combined biophysical and molecular biological methods used to study structure-function.

 Malayev, A., and Nelson, D.J. 1995. Extracellular pH modulates the Ca2+ current activated by depletion of intracellular Ca2+ stores in human macrophages. J. Memb. Biol, 146: 101-111.

 Takeda, M., Nelson, D.J., and Soliven, B. 1995. Calcium signalling in cultured rat oligodendrocytes. Glia, 14: 225-236. 

Nelson, D.J., Tien, X-Y., Xie, W., Brasitus, T.A., Kaetzel, M.A., and Dedman, J.R. 1996. Shrinkage activates a non-selective conductance: Involvement of a Walker-Motif protein and protein kinase C. Am. J. Physiol, 270: C179-C191.

 Xie, W., Kaetzel, M.A., Bruzik, K.S., Dedman, J.R., Shears, S.B., and Nelson, D.J. 1996. Inositol 3,4,5,6-tetrakisphosphate inhibits the calmodulin-dependent protein kinase II-activated chloried conductance in T84 colonic epithelial cells. J. Biol. Chem, 271: 14092-14098.

 Lee, T.E., Philipson, L.H., and Nelson, D.J. 1996. N-type inactivation in the mammalian Shaker K+ channel Kv1.4. J. Memb. Biol, 151: 225-235.

 Naren, A.P., Nelson, D. J., Xie, W., Jovov, B., A., Pevsner, J., Bennett, M.K., Benos, D.J., Quick, M. W., and Kirk, K. L. 1997. Regulation of CFTR chloride channels by syntaxin and Munc18 isoforms. Nature, 390: 302-305.

 Lascola, C. D., Nelson, D. J., Kraig, R. P. 1998. Cytoskeletal actin gates a Cl- channel in neocortical astrocytes. J. Neurosci, 18: 1679-1692.

 Xie, W., Kaetzel, M. A., Bruzik, K. S., Nelson, D. J., and Shears, S. B. 1998. Protein phosphatase-mediated regulation of calcium-dependent chloride conductance in T84 colonic epithelial cells: cross-talk with inositol 3,4,5,6-tetrakisphosphate. J. Physiol. In press.

 Fox, A. P., Hurley, J. H., Nucifora, P. G. P., Nelson, D. J., Ghetti, B., Dlouhy, S., Won, L., and Heller, A. 1998. Altered electrical properties in cerebellar neurons from Weaver heterozygote mice. Exp. Br. Res.

 In press. Holevinsky, K. O., and Nelson, D. J. Direct measurement of particle internalization during phagocytosis in macrophages. In press. Biophys. J. In press.