Abstract List for 2010 meeting

#89: Processing Speed, Neural Efficiency and Working Memory: Their Relationship with MRI Measures in Multiple Sclerosis

Covey, Thomas J ¹; Shucard, Janet L ²; Zivadinov, Robert ³; Shucard, David W ²;

¹Division of Cognitive and Behavioral Neurosciences, Program in Neuroscience; ²Division of Cognitive and Behavioral Neurosciences, ³Buffalo Neuroimaging Analysis Center, SUNY University at Buffalo, School of Medicine and Biomedical Sciences;

In the present study, working memory (WM) performance, processing speed, and neural efficiency/neural noise were assessed in MS patients while undergoing a visual n-back task, and these behavioral measures were related to quantitative MRI measures of structural brain deficits. The results extend our previous findings (e.g., Parmenter et al., 2007) and provide new information about the most sensitive behavioral indices of n-back WM deficits in MS, and about the contribution of neural inefficiency to the slowed processing speed seen in MS patients during WM performance. Even when MS patients performed as well as controls as seen for the lowest WM load condition, they responded more slowly and were less efficient in their speed of responding. The performance accuracy findings indicated that, as hypothesized, the correct match trials, associated with top down processing and executive control, provide a sensitive marker of working memory impairment in MS patients. Further, disruption of executive control during manipulation of information in a high WM load task was associated with global damage that included gray matter; while slowing and inefficient information processing during a demanding WM task was associated primarily with global white matter disruption in MS. Importantly, relationships between processing speed, efficiency, performance and structural MRI measures were seen only during the highest WM load condition, the condition that required the most executive control. These findings and those of others suggest that the MRI/behavioral relationships present exclusively during the highest WM load condition reflect connectivity that involves frontal cortical systems, the site for executive control. Future work will examine variability of EEG Event Related brain activity during the n-back task as a functional marker of neural efficiency and resource allocation in patients and controls.

#96: Novel Association of Interphotoreceptor Binding Protein (IRBP) with Cone Outer Segments

Garlipp, Mary Alice ; Gonzalez-Fernandez, Federico ;

Ross Eye Institute, Department of Ophthalmology, SUNY University at Buffalo, School of Medicine and Biomedical Sciences;

Purpose: IRBP has significant activity in promoting the promotion and release of 11-cis retinal and all-trans retinol respectively. Much attention has been given to the ease with which IRBP can be removed from the IPM by aqueous extraction, there are hints in the literature that not all of the IRBP is available for such extraction. Here, we ask whether IRBP demonstrates a physiologically relevant interaction with structures in the retina.
Methods: Xenopus laevis were selected due to their large photoreceptors, and ability of retinal detachments in light and dark. Full-length Xenopus IRBP (XIRBP) was expressed and purified using a combination of Ni2+ affinity and anion exchange. XIRBP was labeled with Alexa-647 at a 1:1 molar ratio. Anti-Xenopus IRBP serum was used to localize native IRBP. “Wash-resistant” IRBP was detected at sites of exogenous IRBP binding. Neural retinas from light- or dark-adapted animals were detached under HBSS (pH 7.4) and washed 3x in 5mL. Retinas from dark adapted animals were detached and handled under infrared light. Wash-resistant IRBP was detected by indirect immunofluorescence using a secondary antibody conjugated to Alexa-647. In controls, the retina was detached under 4% PFA and not washed. In other experiments, retinas were incubated with XIRBP-647, ovalbumin-647, or unconjugated Alexa-647. Sections were examined in cross-section or whole mounts.
Results: Unwashed retinas matrix staining was diffuse and intense. Wash resistant IRBP mainly labeled photoreceptor outer segments, predominantly cones. Similar results observed in exogenous IRBP experiments. XIRBP-647 staining was observed in cone OS. Ovalbumin-647 and Alexa-647 dye showed minimal fluorescence. Dark adapted retinas in both paradigms showed reduced staining.
Conclusions: IRBP interacts with the cone OS in a light dependent manner. On going experiments are aimed at defining whether the interaction is to the out segment or its matrix sheath.

#73: Ryanodine receptors selectively contribute to taste-evoked calcium signals in mouse taste cells.

Rebello, Michelle R ; Medler, Kathryn F ;

Department of Biological Sciences, SUNY University at Buffalo, College of Arts and Sciences;

Taste buds are sensory end organs that detect chemical substances occurring in foodstuffs and relay the relative information to the brain. Taste stimuli activate distinct signaling pathways in taste receptor cells present in these taste buds. Bitter, umami and sweet taste stimuli activate G-protein coupled receptors (GPCRs) to cause Ca²⁺ release from intracellular stores, which is known to occur in Type II cells via a PLCβ2/IP₃R3 signaling pathway. Sour stimuli depolarize taste cells to cause Ca²⁺ influx through voltage-gated calcium channels (VGCCs), presumably in taste cells with chemical synapses. The transduction pathways of salty stimuli are less well defined. There is also a sub-population of taste cells that express VGCCs and detect bitter taste stimuli but do not express the PLCβ2/IP₃R3 pathway. These cells are termed dual-responsive and appear to express PLCβ3 and IP₃R1. Using immunocytochemistry and calcium imaging, we have shown that ryanodine receptors (RyRs), specifically isoform 1, are expressed in taste cells and it has different physiological roles depending on which signaling pathway is being expressed. In taste cells with VGCCs, we find that RyR1 contributes to the depolarization-induced calcium signal, similar to the functional coupling between RyRs and L- type VGCCs seen in skeletal and cardiac muscle. Our findings reveal an important role for RyRs in the transduction of taste-evoked responses and suggest that calcium signaling in taste cells is more complex than has previously been appreciated.This work was supported by NIDCD DC00635801 and NSF 0917893 to KM.

#82: Identifying Cortical Mechanisms of Tinnitus in Rats

Stolzberg, Daniel ; Salvi, Richard J ;

Center for Hearing and Deafness, SUNY University at Buffalo;

Tinnitus is characterized by the persistent perception of an inescapable phantom sound (e.g. ringing, buzzing) in the absence of any source in the environment. One leading theory of tinnitus implicates aberrant spontaneous activity permitted by cortical disinhibition. In support of this hypothesis, magentoencephalographic recordings from humans with tinnitus reveal decreased energy over the auditory cortex (AC) in the alpha band (8-12 Hz) – a frequency band which positively correlates with inhibitory processes – compared with normal subjects. Similarly, field recordings from the surface of the AC in chronically implanted awake rats showed a significant decrease in alpha energy following a high dose of salicylate; a drug known to reliably induce temporary tinnitus in humans and other animals. In order to investigate how salicylate-induced tinnitus affects inhibitory regulation of cortical activity, extracellular neural activity was recorded simultaneously across the lamina of the AC in anesthetized rats before and following a single systemic salicylate injection (300 mg/kg IP). Following salicylate administration, a decrease in alpha energy was observed near the brain surface and deeper electrodes; however, middle layers of the AC responded with an increase in alpha-band energy. Interestingly, salicylate strongly affected sound-driven responses across the AC lamina in a manner indicative of a cortical disinhibition. In addition, frequency receptive fields of AC neurons expanded and the best frequency of the receptive fields of most neurons shifted towards the behaviorally assessed tinnitus pitch (10-20 kHz). These physiological changes support the hypothesis that cortical disinhibition plays a significant role in the generation of salicylate-induced tinnitus and that tinnitus may emerge from an overrepresentation of cortical neurons tuned to the tinnitus pitch.

#77: Localization of MT₁ Melatonin Receptor Promoter-Driven RFP Protein in the cerebellum of BAC C3H/HeN Transgenic Mice

Adamah-Biassi, Ekue B ; Dubocovich, Margarita L ;

Department of Pharmacology and Toxicology, SUNY University at Buffalo, School of Medicine and Biomedical Sciences;

Melatonin responses are mediated through activation of the MT₁ and MT₂ melatonin receptors. The goal of this study was to determine the cellular distribution of RFP-tagged MT₁ promoter in cerebellar slices from C3H/HeN transgenic mouse expressing Red Fluorescence Protein (RFP) cDNA at the start codon of the MT₁ receptor gene. RFP-tagged MT₁ promoter-containing cells were visualized by direct fluorescence microscopy and by direct immunohistochemistry with RFP antibodies. Free floating cerebellar slices (35 um) from RFP positive and RFP-negative mice were incubated with RFP biotinylated antibody and processed for immunohistochemistry staining and visualization with 3, 3’-Diaminobenzidine (DAB). Expression of RFP tagged MT₁ promoter was found in the granular, the Purkinje and the molecular cell layers. The cell specific expression of RFP tagged MT₁ promoter to Purkinje cells was assessed by double immunohistochemistry using a Purkinje cell specific antibody, calbindin. Cells expressing RFP-tagged MT₁ promoter include Purkinje cells in the Purkinje cells layer, the basket-stellate cells of the molecular layer and the granules cells of the granular layer. We conclude the human and mice cerebellum express MT₁ receptors in the basket-stellate cells and granule cells, while this receptor was also localized in the Purkinje cells of the mouse cerebellum.

#71: Spine density changes in Gonadotropin-Releasing Hormone (GnRH) neurons during the rat estrous cycle

Arroyo, Armando ; Kim, Beom Su ; Garza, Phillip ; Bett, Glenna C L;

Department of Gynecology-Obstetrics and Physiology and Biophysics, SUNY University at Buffalo, School of Medicine and Biomedical Sciences;

Gonadotropin-releasing hormone (GnRH) neurons in the hypothalamus play a critical role in controlling ovulation. Spines are important anatomical structures of GnRH neuron soma and dendrites. Here we determine whether spine density in medial preoptic area (MPOA) GnRH neurons changes across the rat estrous cycle and the effect of estrogen on spines in MPOA GnRH neurons. We measured spine density (# spines/50µm) in MPOA GnRH neuron soma and dendrites during diestrus, proestrus AM, proestrus PM, and estrus in cycling rats (n=5 rats/stage; 5 neurons/rat) using immunohistochemistry and confocal microscopy. We also measured spine density in MPOA GnRH neuron soma and dendrites in OVX rats treated with 17-β estradiol benzonate (n=5 rats/group; 5 neurons/rat) and oil. We found that spine density significantly increased in both GnRH neuron soma and dendrites during proestrus PM. Spine density in both MPOA GnRH neuron soma and dendrites was significantly higher in estrogen compared to oil treated OVX rats. We also determine whether the shape of MPOA GnRH neurons changes across the rat estrous cycle. We found that 62% are unipolar and 30% are bipolar. This proportion did not change across the rat estrous cycle. In conclusion we found that spine density in MPOA GnRH neurons increases during the evening of proestrus during the rat estrous cycle and that estrogen stimulates spine formation in MPOA GnRH neurons. This suggests that spines in GnRH neurons regulate the surge in GnRH secretion that occurs during the evening of proestrus in the rat.

#93: Kinetic Effects of Cross-Linking the LBD Heterodimer Interface in NMDA Receptors

Borschel, William F ; Popescu, Gabriela K ;

Department of Biochemistry, SUNY University at Buffalo, School of Medicine and Biomedical Sciences;

Upon binding the neurotransmitter glutamate and the obligatory co-agonist glycine, NMDA receptors activate by opening a membrane permeable pore or desensitize by switching into a high-affinity non-conducting conformation. Both activation and desensitization require that the agonist-binding clamshell within each subunit closes to engulf the agonists. It has been hypothesized that this movement strains the contacts between agonist-binding domains of GluN1 and GluN2, and rupture of this interface causes receptor desensitization by disengaging agonist-binding from pore-opening. To test the hypothesis of the role of inter-subunit contacts in NMDA receptor gating, we cross-linked the heterodimer interface by introducing cysteine residues at positions predicted to interact across subunits: N521 and L777 of GluN1 and E516 and L780 of GluN2A, respectively. Steady-state single-channel recordings indicated that cross-linked receptors had drastically reduced open probabilities (~150-fold, P_O = 0.0032) due to ~4-fold shorter openings and ~115-fold longer closures (means, SEM): MOT = 1.8 ± 0.2 ms, MCT = 792 ± 213 ms (n = 6; 80,028 events). Surprisingly, the mean duration of closed intervals associated with desensitization remained unaltered (Wt, TauD = 2.7 s; Mut, TauD = 3.0 s). Reduction of the engineered disulfide bonds (10 mM DTT) significantly potentiated single channel currents (means, SEM: P_O = 0.14 ± 0.02) by restoring the mean duration of openings (10.5 ± 0.6 ms) and significantly shortening mean closed durations (80 ± 12 ms), but had no discernible effects on microscopic desensitization (n = 7; 262,396 events). Based on these data, we propose that rearrangements at the heterodimer interface between agonist-binding domains are integral to receptor opening, but not to the desensitizing reaction pathway.

#86: Asymmetric Gating of the Neuromuscular Acetylcholine Receptor Subunits at the Pre-M1 Linker

Bruhova, Iva ; Auerbach, Anthony ;

Department of Physiology and Biophysics, SUNY University at Buffalo, School of Medicine and Biomedical Sciences;

Neuromuscular acetylcholine receptors (AChRs) mediate fast chemical synaptic transmission. Binding of two neurotransmitters at the extracellular domain (ECD) triggers a conformational change causing channel opening at the transmembrane domain (TMD). The ECD/TMD interface is a complex region that links ‘binding’ and ‘gating’. The covalent connection between the ECD and TMD is the pre-M1 linker, a stretch of five residues following β10. In each subunit, this linker has a central Arg (‘R3’), which only in the non-α subunits is flanked by positively-charged residues. Previous studies showed that mutations of R3 in the α subunit alter the gating equilibrium constant and reduce channel expression. We recorded single-channel currents and estimated the gating rate and equilibrium constants of adult mouse AChRs having mutations at the pre-M1 linker and the nearby residue E45 in non-α subunits. In all subunits, mutations of R3 had similar effects as in α. In ε, mutations of the flanking residues and E45 had only small effects, and there was no energy coupling between εE45 and εR3. The non-α R3 residues had Φ values that were similar to that for α. The results suggest that there is a general symmetry between the AChR subunits during gating isomerization in this linker and that the central Arg is involved in expression more so than gating. The energy transfer through the AChR during gating appears to mainly involve position E45, but only in the α subunits.

#81: Differential Expression of Adhesion- and Extracellular Matrix-related Genes among Different Cochlear Partitions and the Inferior Colliculus

Cai, Qunfeng ; Hu, Bohua ;

Center for Hearing and Deafness, SUNY University at Buffalo, College of Arts and Sciences;

The cochlea is a sensory organ consisting of three distinct partitions: the sensory epithelium, the lateral wall, and the modiolus, each playing a distinct role in auditory perception and each responding differently to pathological insults. Investigation into the difference in the gene expression patterns among these structures can provide new insights into molecular mechanisms responsible for normal cochlear function and for cochlear pathogenesis. The current study was designed to profile the expression patterns of adhesion- and extracellular matrix-related genes in the three cochlear partitions and in the inferior colliculus of young Sprague Dawley rats (2-3 months) using a RT-qPCR array technique. The results showed that for all the tissue types, there was a large variation in expression levels among individual genes with the range of the difference being greater than 131,072 folds. In the cochlear sensory epithelia, the highly expressed genes include 12 adhesion-related genes (Ctnnb1, Catna1, Thbs1, Lamb2, Cdh1, Itgb4, Itgav, Sgce, Spp1, Itga3, Cntn1 and Postn) and 8 extracellular matrix-related genes (Sparc, Timp3, Ctgf, Tgfbi, Spock1, Lama2, Mmp11 and Col2a1). A third of examined genes in the lateral wall, the modiolus and the inferior colliculus expressed differently as compared with those expressed in the cochlear sensory epithelia. As expected, the number of genes that differed in the expression level among the three cochlear partitions is less than that showed between the inferior colliculus and the cochlear tissues, suggesting that gene expression patterns are less diverse between cochlear tissues than between cochlear and non-cochlear tissues. Importantly, the study indentified 3 genes (Integrin α3, Periostin, HCAM) in the cochlear sensory epithelium that exhibited higher expression levels than those in other examined tissue types, suggesting that these genes may be involved in sensory cell function. (Supported by NIH R01 DC010154-01A2)

#83: Group I mGluR Activation at the Endbulb of Held

Chanda, Soham ; Xu-Friedman, Matthew A ;

Department of Biological Sciences, SUNY University at Buffalo, College of Arts and Sciences;

Group I metabotropic glutamate receptors (mGluRs) have been shown to play a role in several aspects of neuronal plasticity. We studied the mechanisms and effects of mGluR activation in the mammalian cochlear nucleus. We found that mGluRs are expressed on the postsynaptic bushy cells (BCs) but not in the presynaptic terminals made by auditory nerve (AN) fibers. Activation of mGluRs with dihydroxyphenylglycine (DHPG) caused calcium influx through L-type calcium channels in the BCs. This calcium influx further activated TRP channels causing membrane depolarization. The membrane depolarization was sufficient to enhance the firing probability of BCs in response to AN stimulation and improved the spike timing by reducing the latency and jitter.