Glial cell line-derived neurotrophic factor (GDNF) has emerged as the most potent neuroprotective agent tested in experimental models for the treatment of Parkinson's disease (PD). However, its use is hindered by difficulties in delivery to the brain due to the presence of the blood-brain barrier (BBB). In order to circumvent this problem, we took advantage of the fact that bone marrow stem cell-derived macrophages are able to pass the BBB and home to sites of neuronal degeneration. Here, we report the development of a method for brain delivery of GDNF by genetically modified macrophages. Bone marrow stem cells were transduced ex vivo with lentivirus expressing a GDNF gene driven by a synthetic macrophage-specific promoter and then transplanted into recipient mice. Eight weeks after transplantation, the mice were injected with the neurotoxin, MPTP, for 7 days to induce dopaminergic neurodegeneration. Macrophage-mediated GDNF treatment dramatically ameliorated MPTP-induced degeneration of tyrosine hydroxylase (TH)-positive neurons of the substantia nigra and TH(+) terminals in the striatum, stimulated axon regeneration, and reversed hypoactivity in the open field test. These results indicate that macrophage-mediated GDNF delivery is a promising strategy for developing a neuroprotective therapy for PD.

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Olfactory sensory neurons (OSNs) expressing a specific odorant receptor (OR) gene send axonal projections to specific glomeruli, creating a stereotypic olfactory sensory map. Odorant receptor sequence, G-protein cAMP signaling, and axon guidance molecules have been shown to direct axons of OSNs toward central targets in the olfactory bulb (OB). Although the OR sequence may act as one determinant, our objective was to elucidate the extent by which voltage-dependent activity of postsynaptic projection neurons in the OB centrally influences peripheral development and target destination of OSNs. We bred OR-tagged transgenic mice to homozygosity with mice that had a gene-targeted deletion of the Shaker potassium ion channel (Kv1.3) to elucidate how activity modulates synaptic connections that formulate the sensory map. Here we report that the Kv1.3 ion channel, which is predominantly expressed in mitral cells and whose gene-targeted deletion causes a "super-smeller" phenotype, alters synaptic refinement of axonal projections from OSNs expressing P2, M72, and MOR28 ORs. Absence of Kv1.3 voltage-gated activity caused the formation of small, heterogeneous, and supernumerary glomeruli that failed to undergo neural pruning over development. These changes were accompanied by a significant decrease in the number of P2-, M72-, and MOR28-expressing OSNs, which contained an overexpression of OR protein and G-protein G olf in the cilia of the olfactory epithelium. These findings suggest that voltage-gated activity of projection neurons is essential to refine primary olfactory projections and that it regulates proper expression of the transduction machinery at the periphery. Indexing termsolfactory coding; axon targeting; olfaction; odorant receptor; Kv1.3The development of precise connectivity and maintenance of an olfactory sensory map is thought to rely upon guidance molecules that are recognized by receptors on olfactory sensory neurons (OSNs) or that are regulated by G-protein-mediated cAMP signals (Bozza et al., 2002;St John et al., 2002;Imai et al., 2006; Lattermann et al., 2007;Sweeney et al., 2007). OSNs expressing a given olfactory receptor are primarily randomly dispersed within one of four zones in the main olfactory epithelium (Ressler et al., 1993;Vassar et al., 1993;Mombaerts et al., 1996) but send their axonal projections to spatially conserved glomeruli within the olfactory bulb (Mombaerts, 1996;Wang et al., 1998;Tsuboi et al., 1999). Thus the olfactory bulb (OB) serves as a two- NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript dimensional map of receptor activation to provide the neural code to discriminate olfactory sensory information (Yu et al., 2004). Odorant quality is thus encoded by a specific combination of activated glomeruli (Mori et al., 1999;Strotmann et al., 2000), and the formation of glomeruli depends on contextual sorting of axons expressing like OR proteins (Belluscio et al., 2002;Mombaerts, 2006).If contextual cues from adjacent homotypic interactions initiate...

Kv subfamily member 1.3 (Kv1.3), a member of the Shaker family of potassium channels, has been found to play diverse roles in immunity, metabolism, insulin resistance, sensory discrimination, and axonal targeting in addition to its traditional role in the stabilization of the resting potential. We demonstrate that the neurotrophin B receptor (TrkB) causes an upregulation of Kv1.3 ion channel protein expression in the absence of the preferred ligand for the receptor (brain-derived neurotrophic factor; BDNF) and oppositely downregulates levels of Kv subfamily member 1.5. Although the effect occurs in the absence of the ligand, Kv1.3 upregulation by TrkB is dependent upon the catalytic domain of the TrkB kinase as well as tyrosine (Y) residues in the N and C terminus of the Kv1.3 channel. Using pulse-chase experiments we find that TrkB alters the half-life residence of the channel by approximately 2x and allows it to sustain activity as reflected in an increased current magnitude without alteration of kinetic properties. TrkB and Kv1.3 co-immunoprecipitate from tissue preparations of the mouse olfactory bulb and olfactory cortex, and by immunocytochemical approaches, are found to be co-localized in the glomerular, mitral cell, and internal plexiform layers of the olfactory bulb. These data suggest that Kv1.3 is not only modulated by direct phosphorylation in the presence of BDNF-activated TrkB kinase, but also may be fine tuned via regulation of surface expression while in the proximity of neurotrophic factor receptors. Given the variability of TrkB expression during development, regeneration, or neuronal activation, modulation of surface expression and turnover of Kv channels could significantly impact neuronal excitability, distinct from that of tyrosine kinase phosphorylation.

The voltage-gated potassium channel, Kv1.3, contributes a large proportion of the current in mitral cell neurons of the olfactory bulb where it assists to time the firing patterns of action potentials as spike clusters that are important for odorant detection. Gene-targeted deletion of the Kv1.3 channel, produces a “super-smeller” phenotype, whereby mice are additionally resistant to diet- and genetically-induced obesity. As assessed via an electrophysiological slice preparation of the olfactory bulb, Kv1.3 is modulated via energetically important molecules – such as insulin and glucose – contributing to the body’s metabolic response to fat intake. We discuss a biophysical characterization of modulated synaptic communication in the slice following acute glucose and insulin stimulation, chronic elevation of insulin in mice that are in a conscious state, and induction of diet-induced obesity. We have discovered that Kv1.3 contributes an unusual nonconducting role – the detection of metabolic state.