Faculty Bibliography

Cerebellar slices from 11-13 day old rat brains were cultured for 2 weeks in Minimum Essential Medium with standard 5 mM KCL. At 14 day in vitro (DIV14), calbindin immunoreactivity and Nissl staining revealed many surviving Purkinje cells (PCs) but few granule cells. Ultrastructually, the neuropil was composed mostly of PC dendrites with small and long thin spines but few presynaptic boutons, since granule cell axons (GC parallel fibers) and boutons were negligible. Three experimental groups, exposed to increased K+ levels (10, 20 and 30 mM KCL) between DIV 7-14, had increased number of GCs but fewer PCs with shortened dendritic arbors. EM analysis revealed prominent long segments of parallel fibers and enlarged boutons with abundant synaptic vesicles. Bouton size increased consecutively with each higher concentration of KCL. Some large boutons were opposed to PC plasma membrane and formed multiple synaptic contacts with spines emerging from PC somata and various sized dendrites. Occasionally, 20 or more spine heads contacted each giant bouton. On the other hand, PCs had atrophy with cytological changes in mitochondria and swelling of PC dendrites with stacks of ER. This study shows that above normal K+ enhances GC axonal growth to form giant boutons and supports previous findings that GCs in dissociated cell cultures survive best with 25-30 mM KCL

Oxidative stress is widely considered to be involved in the pathogenesis of various neurological disorders to include Alzheimer's disease. This stress is thought to include, in addition to cell death, neuronal dysfunctions such as neurotransmission deficits. Here, we examined H2O2-induced alterations of electrophysiological properties of Purkinje cell in acute cerebellar slices, and assessed the effects of several neuroprotective agents.Electrophysiological and fura-2 fluorescence imaging techniques were used to determine intercellular calcium concentration ((Ca2+)i). H2O2 (100 muM) reversibly reduced both holding current and EPSC amplitude in voltage clamped neurons, and produced, in addition, (Ca2+)i elevation. Both the antioxidant 2-mercaptoethanol and the neuroprotective/neurotrophic agent T-817 (1-\{3-(2-(1-benzothiophen-5-yl)ethoxy)propyl\}-3-azethidinol maleate), partially inhibit these physiological changes. We are presently investigating the mechanism for reactive oxygen spices alteration of neuronal properties, mitocondrial function, (Ca2+)i mobilization, and cell viability, in the presence of the T-817

Intracellular recording and in vitro high-speed voltage-sensitive dye imaging were combined to investigate the role of electrotonic coupling in the generation and distribution of subthreshold oscillations in the inferior olivary (IO) nucleus. Brainstem slices from 2-3 week old rats were stained with a voltage-sensitive fluorescence dye RH-414 (Molecular Probes) and imaged with a fast CCD camera (Fujix HRDeltaron 1700; 128 x 128 pixels of spatial resolution and 4.8 ms of temporal resolution). Comparison of spatio-temporal profiles of subthreshold IO oscillations in control conditions and in the presence a specific gap junctional blocker (18-beta glycyrrhetinic acid) was implemented. In control conditions, spontaneous IO oscillations emanated from multiple clusters of coherently oscillating neurons. Addition of 18-beta glycyrrhetinic acid had a blocking effect such clusters. However, single-cell oscillations were still observed in concurrently obtained intracellular recordings from individual IO cells. Small but significant differences in the amplitude and frequency of the 'uncoupled' and control neuronal oscillations were detected. In addition, the cells had higher resistance (10-20% increase) and lower capacitance (20-30% decrease) in the presence of the blocker. We conclude that gap junctions are not necessary for generating subthreshold IO oscillations but are required for oscillatory synchronization and the clustering of coherent oscillatory activity in the inferior olivary nucleus

Thalamic relay cells occupy a pivotal position in cerebral architecture, and characterizing the manner in which they integrate ascending and descending input is a requisite for understanding cortical function. It was demonstrated by McCormick and Huguenard in the early 1990's that the relay and low threshold burst modes of intrinsic response can be readily modeled using a single somatic compartment with an appropriate set of voltage and calcium concentration-gated currents. However, the study of synaptic integration in model cells requires simulations incorporating the dendritic tree. Destexhe, Huguenard and coworkers have decribed a relay cell model including a dendritic tree (Destexhe et al 1998) which concluded a predominantly distal T-channel density distribution, primarily constrained by recordings from acutely dissociated cells. There is however another intrinsic electrophysiological characteristic of relay cells which models have not yet accounted for: at holding potentials near spike threshold, relay cells produce a waxing and waning subthreshold oscillation observable at the soma (Pedroarena and Llinas 1996). Here we present the development of a new model of the thalamic relay cell, guided by the simultaneous constraints that it must produce the relay and low threshold burst modes which characterize these cells, as well as the ripple occurring at near-threshold holding potentials. We arrive at a model cell which is capable of the production isolated fast Ca2+ spikes in distal branch segments, driven by a rapidly inactivating high threshold channel. The model reproduces the low threshold spike behavior of the relay cell without requiring high T current density in the distal dendritic segments, and thus presents an alternative picture of the dendritic tree of relay cells, in which fast high threshold Ca2+ events occur distally, and slower T channel-driven spikes are primarily proximally driven. If borne out by direct experimental evidence, this model would have implications for the integration of descending and ascending inputs

Calcium mobilization in cerebellar Purkinje cell (PC) of CD38 knockout mouse was imaged using two photon microscopy. CD38 is a homolog of ADP-ribosyl cyclase which activates Cyclic ADP-ribose, an endogenous Calcium Induced Calcium Release (CICR)regulator (Lee, 2001 ). In the CD38 knockout mouse (CD38(-/-)), which showed no phenotypic motor abnormalities, whole cerebellar cyclic ADP-ribose (cADPR) was reduced to a quarter of that in a wild control (CD38(+/+)). In addition caffeine treated CD38(-/-) PC dendrite showed a markedly reduced calcium response to a test depolarizing pulse most probably related to the reduced CICR. Cerebellar long-term depression (LTD) induced by the parallel fiber (PF) -climbing fiber (CF) conjunctional stimulation was absent in CD38(-/-) PC. However, during very prolonged direct depolarization of Purkinje cell somata, or when the test (PF) stimulation was quite close to the cell body during direct somatic depolarization, LTD could be induced in CD38(-/-) PC by the PF/depolarizing pulse conjunction. Overall, the CD38/cADPR system seems to be sensing/regulating intradendritic calcium levels. This seems to be an important modulator of the integrative properties of Purkinje cell activity. It seems clear that if (Ca)i becomes high enough to be deleterious to neuronal viability such system must be down regulated via LTD as a neuroprotective response to reduce further calcium increase. Immuno-staining study with a polyclonal CD38 antibody is presently in progress which is expected to find different levels of CD38 on the Purkinje cell soma, dendrites and spines of wild type mouse (CD38(+/+)) vs CD38 knockout cerebella

One of the essential aspects of the neuronal organization in the global function of the brain is the rich thalamocortical interconnectivity and very particularly the reciprocal nature of this circuit. Also, the interaction between the systems specific thalamic and unspecific at cortical level suggests that the thalamus, more than a simple floodgate for the brain, represents an epicentre by means of which all the cortical areas can communicate to each other in isochronic way with independence of the transcortical distance. The objectives of this article are to explore: 1) the proposal that the temporary coincidence, to great scale, of the activity specific thalamic and unspecific generates the functional states that characterize the human knowledge; and 2) the possible relationship between the thalamocortical dysrhythmia and some neuropsychiatric illnesses

The sources of the neuromagnetic fields recorded in Magnetoencephalography (MEG) are distributed compactly as sparse clusters of high current density in functionally interactive cortical and subcortical neural networks. Recursive minimum norm algorithms (e.g., FOCUSS) iteratively sharpen and eventually prune an initial distributed solution by down-weighting the contribution of low-amplitude dipoles. Most nonzero variables are not needed to explain the data and can be eliminated by iteratively adding a null space vector to the previous solution that minimizes a diversity measure and guarantees feasibility. Computer simulations with randomly selected source configurations of increasing order and with sparse extended sources were carried out to quantify the spatial resolution of recursive minimum-norm algorithms with several spatial error metrics. Different a priori weights and diversity measures were tested. Results demonstrate that stable super-resolution for high-order source models can only be achieved by using an optimal a priori weight matrix (full column normalization), followed by a sequence of iterations which minimize an optimal p-norm-like diversity measure (p=0.9-1). Further improvement is achieved by directly minimizing a cost of diversity and discrepancy at each iteration with a nonlinear 1-dimensional search of the optimal null vector length, and by simultaneously solving multiple measurement vectors from a time interval. Simulations with non-gaussian activation functions show that Independent Component Analysis (ICA) and the new FOCUSS-learning algorithms can dramatically increase the number of recovered sources. Finally, visually evoked and spontaneous signals are analyzed with these algorithms. Results demonstrate the importance of cortico-thalamo-cortical loops in global brain function

Clearly, more clinical experience must be amassed to define in detail the possibilities of this surgical approach in disabling neuropsychiatric disorders. We propose, however, that the evidence for benign and efficient surgical intervention against the neuropsychiatric TCD syndrome is already compelling. The potential appearance of strong postoperative reactive manifestations requires a close association between surgery and psychotherapy, with the latter providing support for the integration of the new situation as well as the resolution of old unresolved issues