Faculty Bibliography

The increased availability of transgenic mouse models for studying human diseases is shifting the focus of many laboratories from in vitro to in vivo assays. The purpose of this chapter is to provide investigators with methods that will allow them to obtain well-preserved mouse brain sections to be stained with the standard histological dyes for amyloid, Congo red and thio-flavin-S. These sections can as well be used for immunohistological procedures that allow detection of amyloid-beta plaques as well as pre-amyloid deposits

Transgenic mice are used increasingly to model brain amyloidosis, mimicking the pathogenic processes involved in Alzheimer's disease (AD). In this chapter, a strategy is described that has been successfully used to map amyloid deposits in transgenic mouse models of AD with magnetic resonance imaging (MRI), utilizing molecular targeting vectors labeled with MRI contrast agents to enhance selectively the signal from amyloid plaques. To obtain sufficient spatial resolution for effective and sensitive mouse brain imaging, magnetic fields of 7-Tesla (T) or more are required. These are higher than the 1.5-T field strength routinely used for human brain imaging. The higher magnetic fields affect contrast agent efficiency, and determine the choice of pulse sequence parameters for in vivo MRI, all addressed in this chapter. Ex vivo imaging is also described as an important step to test and optimize protocols prior to in vivo studies. The experimental setup required for mouse brain imaging is explained in detail, including anesthesia, immobilization of the mouse head to reduce motion artifacts, and anatomical landmarks to use for the slice alignment procedure to improve image co-registration during longitudinal studies, and for subsequent matching of MRI with histology

In recent years major outbreaks of prion disease linked to oral exposure of the prion agent have occurred in animal and human populations. These disorders are associated with a conformational change of a normal protein, PrP(C) (prion protein cellular), to a toxic and infectious form, PrP(Sc) (prion protein scrapie). None of the prionoses currently have an effective treatment. A limited number of active immunization approaches have been shown to slightly prolong the incubation period of prion infection. Active immunization in wild-type animals is hampered by auto-tolerance to PrP and potential toxicity. Here we report that mucosal vaccination with an attenuated Salmonella vaccine strain expressing the mouse PrP, is effective at overcoming tolerance to PrP and leads to a significant delay or prevention of prion disease in mice later exposed orally to the 139A scrapie strain. This mucosal vaccine induced gut anti-PrP immunoglobulin (Ig)A and systemic anti-PrP IgG. No toxicity was evident with this vaccination approach. This promising finding suggests that mucosal vaccination may be a useful method for overcoming tolerance to PrP and preventing prion infection among animal and potentially human populations at risk

We used the juxtacellular recording and labeling technique of Pinault (1996) in the uvula/nodulus of the ketamine anesthetized rat in an attempt to link different patterns of spontaneous activity with different types of morphologically identified cerebellar cortical interneurons. Cells displaying a somewhat irregular, syncopated cadence of spontaneous activity averaging 4-10 Hz could, upon successful entrainment and visualization, be morphologically identified as Golgi cells. Spontaneously firing cells with a highly or fairly regular firing rate of 10-35 Hz turned out to be unipolar brush cells. We also found indications that other types of cerebellar cortical neurons might also be distinguished on the basis of the characteristics of their spontaneous firing. Comparison of the interspike interval histograms of spontaneous activity obtained in the anaesthetized rat with those obtained in the awake rabbit points to a way whereby the behaviorally related modulation of specific types of interneurons can be studied. In particular, the spontaneous activity signatures of Golgi cells and unipolar brush cells anatomically identified in the uvula/nodulus of the anaesthetized rat are remarkably similar to the spontaneous activity patterns of some units we have recorded in the flocculus of the awake rabbit. The spontaneous activity patterns of at least some types of cerebellar interneurons clearly have the potential to serve as identifying signatures in behaving animals

Postural related changes in cerebral hemodynamics and hydrodynamics were studied using Magnetic Resonance Imaging (MRI) measurements of cerebral blood flow and cerebrospinal fluid (CSF) flow dynamics. Ten healthy volunteers (mean age 29 +/- 7) were studied in supine and upright (sitting) postures. A Cine phase-contrast MRI technique was used to image the pulsatile blood flow to the brain, the venous outflow through the internal jugular, epidural, and vertebral veins, and the bi-directional CSF flow between the cranium and the spinal canal. Previously published analyses were applied to calculate and compare total cerebral blood flow (TCBF), intracranial compliance and pressure in both postures. A lower (12%) mean TCBF was measured in the upright position compared to supine position. A considerable smaller amount of CSF flow between the cranium and the spinal canal (58%), a much larger intracranial compliance (a 2.8-fold increase), and a corresponding decrease in the MRI-derived ICP were also measured in the sitting position. These changes suggest that the increased cerebrovascular and intracranial compliances in the upright posture are primarily due to reduced amounts of blood and CSF residing in their respective intracranial compartments in the upright position. This work demonstrates the ability to quantify neurophysiologic parameters associated with regulation of cerebral hemodynamics and hydrodynamics from dynamic MR imaging of blood and CSF flows.

Cerebral blood flow and ICP are important neurophysiologic parameters known to be affected by pathology and by trauma. Limited data on the relationship between these parameters following head trauma is inconsistent with regard to whether these parameters are correlated. Data on the relationship between these parameters in the healthy state is not readily available due to a lack of noninvasive means to measure these important parameters. A recently developed noninvasive MRI-based method for simultaneous measurement of total cerebral blood flow and intracranial pressure was applied to establish the relationship between ICP and TCBF values in healthy subjects. Seventy-one simultaneous measurements of CBF and ICP were obtained from 23 healthy young adults. These results demonstrated that CBF values span over a much narrower range compared with ICP. The relationship between the inter-individual CBF and ICP measurements suggest that in the healthy state and in rest these parameters are not correlated.

Cochlear implants are electrical prostheses that partially replace the functions of the human ear. They bypass normal hearing operation to directly simulate the auditory nerve with electric current. The input acoustic signal passes through a filter bank and the output of each filter modulates the energy of a stimulation waveform delivered to a different intra-cochlear electrode. This approach attempts to mimic the signal processing that takes place in a normal ear. When fitting a cochlear implant to a patient who has lost his hearing after learning language, one important problem is how to optimize the frequency range of the filter bank This optimization seeks a tradeoff between maximum speech perception and the patient\\\\\\\'s subjective preference. Unfortunately, currently available tools to change the frequency-to-electrode mapping (i.e., the frequencies of the filter bank) are cumbersome to use. In a previous project we developed a real time speech processor for the Nucleus-22 and Nucleus-24 cochlear implants, based on a common PC and additional hardware drivers. The present project involves the development of a similar system that is adjustable in real time. In other words, the patient can modify the frequency-to-electrode map using computer keystrokes, and a visual representation of the frequency range employed by the filter bank is displayed on the monitor. The patient adjusts the frequency range interactively and selects the preferred setting in a much faster way than can be accomplished with commercially available hardware. If successful, this approach may be implemented in the next generation of hardware used to program cochlear implants in the clinic

The olivo-cerebellar network is a key neuronal circuit that provides high-level motor control in the vertebrate CNS. Functionally, its network dynamics is organized around the oscillatory membrane potential properties of inferior olive (IO) neurons and their electrotonic connectivity. Because IO action potentials are generated at the peaks of the quasisinusoidal membrane potential oscillations, their temporal firing properties are defined by the IO rhythmicity. Excitatory inputs to these neurons can produce oscillatory phase shifts without modifying the amplitude or frequency of the oscillations, allowing well defined time-shift modulation of action potential generation. Moreover, the resulting phase is defined only by the amplitude and duration of the reset stimulus and is independent of the original oscillatory phase when the stimulus was delivered. This reset property, henceforth referred to as selfreferential phase reset, results in the generation of organized clusters of electrically coupled cells that oscillate in phase and are controlled by inhibitory feedback loops through the cerebellar nuclei and the cerebellar cortex. These clusters provide a dynamical representation of arbitrary motor intention patterns that are further mapped to the motor execution system. Being supplied with sensory inputs, the olivo-cerebellar network is capable of rearranging the clusters during the process of movement execution. Accordingly, the phase of the IO oscillators can be rapidly reset to a desired phase independently of the history of phase evolution. The goal of this article is to show how this selfreferential phase reset may be implemented into a motor control system by using a biologically based mathematical model.

Synaptotagmin (Syt) I, a ubiquitous synaptic vesicle protein, comprises a transmembrane region and two C2 domains. The C2 domains, which have been shown to be essential for both synaptic vesicle exocytosis and endocytosis, are also seen as the Ca(2+) sensors in synaptic vesicular release. In a previous study, we reported that a polyclonal antibody raised against the squid (Loligo pealei) Syt I C2B domain, while inhibiting vesicular endocytosis, was synaptic release neutral at the squid giant synapse. Recent reports concerning the C2B requirements for synaptic release prompted us to readdress the role of C2B in squid giant synapse function. Presynaptic injection of another anti-Syt I-C2B antibody (using recombinant whole C2B domain expressed in mammalian cell culture as an antigen) into the presynaptic terminal reproduced our previous results, i.e., reduction of vesicular endocytosis without affecting synaptic release. This set of results addresses the issue of the geometrical arrangement of the Ca(2+) sensor, allowing the C2B domain antibody to restrict Ca(2+)-dependent C2B self-oligomerization without modifying the Ca(2+)-dependent release process

Inferior olivary (IO) neurons display spontaneous oscillatory activity, yet the importance of these oscillations for shaping the responses of this system to its afferents is uncertain. We used multiple electrode recording of crus 2a Purkinje cell complex spikes (CSs) in ketamine-xylazine-anesthetized rats to investigate olivocerebellar responses to activation of motor cortico-olivary pathways. Trains of electrical stimuli were applied to the motor cortex at frequencies between 4 and 30 Hz. Various frequency-response curves were observed, with the most common types being unimodal with a maximum at 9.5 +/- 2.3 Hz and bimodal with peaks at 8.9 +/- 1.0 and 15.1 +/- 1.3 Hz. To determine whether IO oscillatory properties underlie the resonance peaks in the frequency-response curves, apamin and charybdotoxin were injected into the IO. These toxins, which weaken and enhance spontaneous IO oscillations, respectively, had corresponding effects on the sharpness of resonance peaks. Next, the variation of CS entrainment patterns with frequency was investigated to characterize the nature of the IO oscillator. Low-frequency (4 Hz) stimulation was relatively ineffective in entraining CS activity. Between 4 and 30 Hz, two predominant entrainment patterns emerged. For low-frequency (4-6 Hz) and high-frequency (17-30 Hz) ranges, a 1:2 entrainment dominated, whereas in the intermediate range (6-17 Hz), 1:1 entrainment was most prevalent. These results indicate that IO neurons respond as nonlinear oscillators to afferent signals