Faculty Bibliography

Background- The specialized cardiac conduction system (CCS) expresses a unique complement of ion channels that confer a specific electrophysiological profile. ATP-sensitive potassium (K(ATP)) channels in these myocytes have not been systemically investigated. Methods and Results- We recorded K(ATP) channels in isolated CCS myocytes using Cntn2-EGFP reporter mice. The CCS K(ATP) channels were less sensitive to inhibitory cytosolic ATP compared with ventricular channels and more strongly activated by MgADP. They also had a smaller slope conductance. The 2 types of channels had similar intraburst open and closed times, but the CCS K(ATP) channel had a prolonged interburst closed time. CCS K(ATP) channels were strongly activated by diazoxide and less by levcromakalim, whereas the ventricular K(ATP) channel had a reverse pharmacological profile. CCS myocytes express elevated levels of Kir6.1 but reduced Kir6.2 and SUR2A mRNA compared with ventricular myocytes (SUR1 expression was negligible). SUR2B mRNA expression was higher in CCS myocytes relative to SUR2A. Canine Purkinje fibers expressed higher levels of Kir6.1 and SUR2B protein relative to the ventricle. Numeric simulation predicts a high sensitivity of the Purkinje action potential to changes in ATP:ADP ratio. Cardiac conduction time was prolonged by low-flow ischemia in isolated, perfused mouse hearts, which was prevented by glibenclamide. Conclusions- These data imply a differential electrophysiological response (and possible contribution to arrhythmias) of the ventricular CCS to K(ATP) channel opening during periods of ischemia

PURPOSE: Extended warm ischemia during partial nephrectomy can lead to considerable renal injury. Using a rat model of renal ischemia we examined the ability of a unique renoprotective cocktail to ameliorate warm ischemia-reperfusion injury. MATERIALS AND METHODS: A warm renal ischemia model was developed using 60 Sprague-Dawley(R) rats. The left renal artery was clamped for 40 minutes, followed by 48 hours of reperfusion. A renoprotective cocktail of a mixture of specific growth factors, mitochondria protecting biochemicals and Manganese-Porphyrin (MnTnHex-2-PyP(5+)) was given intramuscularly at -24, 0 and 24 hours after surgery. At 48 hours the 2 kidneys were harvested and examined with hematoxylin and eosin, and periodic acid-Schiff stains. Protein and gene expression were also analyzed to determine ischemia markers and the antioxidant response. RESULTS: Compared to ischemic controls, kidneys treated with the renoprotective cocktail showed significant reversal of morphological changes and a significant decrease in the specific ischemic markers lipocalin-2, mucin-1 and galectin-3. Quantitative reverse transcriptase-polymerase chain reaction revealed up-regulation of several antioxidant genes in treated animals. CONCLUSIONS: According to histopathological and several molecular measures our unique renoprotective cocktail mitigated ischemia-reperfusion injury

Several lines of research have now suggested the controversial hypothesis that the central noradrenergic system acts to exacerbate depression as opposed to having an antidepressant function. If correct, lesions of this system should increase resistance to depression, which has been partially but weakly supported by previous studies. The present study reexamined this question using two more recent methods to lesion noradrenergic neurons in mice: intraventricular (ivt) administration of either the noradrenergic neurotoxin, DSP4, or of a dopamine-beta-hydroxylase-saporin immunotoxin (DBH-SAP ITX) prepared for mice. Both agents given 2weeks prior were found to significantly increase resistance to depressive behavior in several tests including acute and repeated forced swims, tail suspension and endotoxin-induced anhedonia. Both agents also increased locomotor activity in the open field. Cell counts of brainstem monoaminergic neurons, however, showed that both methods produced only partial lesions of the locus coeruleus and also affected the dorsal raphe or ventral tegmental area. Both the cell damage and the antidepressant and hyperactive effects of ivt DSP4 were prevented by a prior i.p. injection of the NE uptake blocker, reboxetine. The results are seen to be consistent with recent pharmacological experiments showing that noradrenergic and serotonergic systems function to inhibit active behavior. Comparison with previous studies utilizing more complete and selective LC lesions suggest that mouse strain, lesion size or involvement of multiple neuronal systems are critical variables in the behavioral and affective effects of monoaminergic lesions and that antidepressant effects and hyperactivity may be more likely to occur if lesions are partial and/or involve multiple monoaminergic systems

Neuromuscular synapse formation depends upon coordinated interactions between motor neurons and muscle fibers, leading to the formation of a highly specialized postsynaptic membrane and a highly differentiated nerve terminal. Synapse formation begins as motor axons approach muscles that are prepatterned in the prospective synaptic region in a manner that depends upon Lrp4, a member of the LDL receptor family, and muscle-specific kinase (MuSK), a receptor tyrosine kinase. Motor axons supply Agrin, which binds Lrp4 and stimulates further MuSK phosphorylation, stabilizing nascent synapses. How Agrin binds Lrp4 and stimulates MuSK kinase activity is poorly understood. Here, we demonstrate that Agrin binds to the N-terminal region of Lrp4, including a subset of the LDLa repeats and the first of four beta-propeller domains, which promotes association between Lrp4 and MuSK and stimulates MuSK kinase activity. In addition, we show that Agrin stimulates the formation of a functional complex between Lrp4 and MuSK on the surface of myotubes in the absence of the transmembrane and intracellular domains of Lrp4. Further, we demonstrate that the first Ig-like domain in MuSK, which shares homology with the NGF-binding region in Tropomyosin Receptor Kinase (TrKA), is required for MuSK to bind Lrp4. These findings suggest that Lrp4 is a cis-acting ligand for MuSK, whereas Agrin functions as an allosteric and paracrine regulator to promote association between Lrp4 and MuSK

Learning to adapt to a complex and fluctuating environment requires the ability to adjust neural representations of sensory stimuli. Through pattern completion processes, cortical networks can reconstruct familiar patterns from degraded input patterns, whereas pattern separation processes allow discrimination of even highly overlapping inputs. Here we show that the balance between pattern separation and completion is experience dependent. Rats given extensive training with overlapping complex odorant mixtures showed improved behavioral discrimination ability and enhanced piriform cortical ensemble pattern separation. In contrast, behavioral training to disregard normally detectable differences between overlapping mixtures resulted in impaired piriform cortical ensemble pattern separation (enhanced pattern completion) and impaired discrimination. This bidirectional effect was not found in the olfactory bulb; it may be due to plasticity within olfactory cortex itself. Thus pattern recognition, and the balance between pattern separation and completion, is highly malleable on the basis of task demands and occurs in concert with changes in perceptual performance

Natural odors, generally composed of many monomolecular components, are analyzed by peripheral receptors into component features and translated into spatiotemporal patterns of neural activity in the olfactory bulb. Here, we will discuss the role of the olfactory cortex in the recognition, separation and completion of those odor-evoked patterns, and how these processes contribute to odor perception. Recent findings regarding the neural architecture, physiology, and plasticity of the olfactory cortex, principally the piriform cortex, will be described in the context of how this paleocortical structure creates odor objects

PURPOSE: Problems in management of oral cancers or precancers include identification of patients at risk for metastasis, tumor recurrence, and second primary tumors or risk for progression of precancers (dysplasia) to cancer. Thus, the objective of this study was to clarify the role of genomic aberrations in oral cancer progression and metastasis. EXPERIMENTAL DESIGN: The spectrum of copy number alterations in oral dysplasia and squamous cell carcinomas (SCC) was determined by array comparative genomic hybridization. Associations with clinical characteristics were studied and results confirmed in an independent cohort. RESULTS: The presence of one or more of the chromosomal aberrations +3q24-qter, -8pter-p23.1, +8q12-q24.2, and +20 distinguishes a major subgroup (70%-80% of lesions, termed 3q8pq20 subtype) from the remainder (20%-30% of lesions, non-3q8pq20). The 3q8pq20 subtype is associated with chromosomal instability and differential methylation in the most chromosomally unstable tumors. The two subtypes differ significantly in clinical outcome with risk for cervical (neck) lymph node metastasis almost exclusively associated with the 3q8pq20 subtype in two independent oral SCC cohorts. CONCLUSIONS: Two subtypes of oral lesions indicative of at least two pathways for oral cancer development were distinguished that differ in chromosomal instability and risk for metastasis, suggesting that +3q,-8p, +8q, and +20 constitute a biomarker with clinical utility for identifying patients at risk for metastasis. Moreover, although increased numbers of genomic alterations can be harbingers of progression to cancer, dysplastic lesions lacking copy number changes cannot be considered benign as they are potential precursors to non-3q8pq20 locally invasive, yet not metastatic oral SCC.

Patterning the Drosophila retina for color vision relies on postmitotic specification of photoreceptor subtypes. R8 photoreceptors express one of two light-sensing Rhodopsins, Rh5 or Rh6. This fate decision involves a bistable feedback loop between Melted, a PH-domain protein, and Warts, a kinase in the Hippo growth pathway. Here, we show that a subset of the Hippo pathway-Merlin, Kibra, and Lethal(2)giant larvae (Lgl), but not Expanded or Fat-is required for Warts expression and activity in R8 to specify Rh6 fate. Melted represses warts transcription to disrupt Hippo pathway activity and specify Rh5 fate. Therefore, R8 Hippo signaling exhibits ON-or-OFF regulation, promoting mutually exclusive fates. Furthermore, Merlin and Lgl are continuously required to maintain R8 neuronal subtypes. These results reveal roles for Merlin, Kibra, and Lgl in neuronal specification and maintenance and show that the Hippo pathway is reimplemented for sensory neuron fate by combining canonical and noncanonical regulatory steps.

Signaling through cyclic AMP (cAMP) has been implicated in the regulation of Schwann cell (SC) proliferation and differentiation. In quiescent SCs, elevation of cAMP promotes the expression of proteins associated with myelination such as Krox-20 and P0, and downregulation of markers associated with the non-myelinating SC phenotype. We have previously shown that the motor protein myosin II is required for the establishment of normal SC-axon interactions, differentiation and myelination, however, the mechanisms behind these effects are unknown. Here we report that the levels and activity of myosin light chain kinase (MLCK), an enzyme that regulates MLC phosphorylation in non-muscle cells, are dramatically downregulated in SCs after cAMP treatment, in a similar pattern to that of c-Jun, a known inhibitor of myelination. Knockdown of MLCK in SCs mimics the effect of cAMP elevation, inducing plasma membrane expansion and expression of Krox-20 and myelin proteins. Despite activation of myelin gene transcription these cells fail to make compact myelin when placed in contact with axons. Our data indicate that myosin II activity is differentially regulated at various stages during myelination and that in the absence of MLCK the processes of SC differentiation and compact myelin assembly are uncoupled.