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Open Access Publications from the University of California

Scholarly Works (23 results)

  • Thesis
  • Peer Reviewed
UCLA Electronic Theses and Dissertations (2014)

&gamma-aminobtutyric acid (GABA) is a transmitter molecule found in several organs and virtually all organisms. In the vertebrate central nervous system it is the major inhibitory neurotransmitter. Here GABA released by GABAergic neurons binds to GABA receptors and regulates neuronal excitability, network synchrony and neuronal development. GABAA receptors (GABAARs) are highly conserved ligand-gated ion channels permeable to Cl- and HCO3-, and upon binding GABA they generate currents that can be functionally distinguished in phasic or tonic. Physiology of tonic GABAergic conductances is of particular interest as these exercise powerful constrain on neuronal excitability and gain. Tonic inhibition is mediated by GABAARs containing &delta or &alpha5 subunits localized outside of the synaptic cleft and activated by ambient GABA. Object of this dissertation is plasticity of &delta- GABAARs, as their function and distinct pharmacology makes them relevant to many human diseases. &delta-GABAARs are naturally insensitive to benzodiazepines while uniquely sensitive to low concentrations of neurosteroids, which act on them as positive allosteric modulators. These endogenous steroids are the neuroactive form of progesterone, cortisol and testosterone. They are locally synthetized in both neurons and glia and their brain concentration varies in parallel with oscillations in plasma precursors. The present work shows how during times of altered neurosteroid production &delta-GABAARs expression homeostatically changes with functional consequences on neuronal network functioning. In particular, plasticity of &delta-GABAARs during pregnancy and the postpartum affects hippocampal excitability and &gamma oscillations frequency in vitro. Moreover, over the ovarian cycle, plasticity of &delta-GABAARs on interneurons is necessary for fluctuations in &gamma oscillations amplitude in vivo. More and more evidence suggests an involvement of &delta-GABAARs in different kinds of neurological and psychiatric disorders, such as epilepsies, postpartum depression, pre-menstrual dysphoric disorder, and schizophrenia. Interestingly a convenient therapeutic strategy may be available, given the anatomical distribution of these receptors. In fact, &delta-GABAARs expressed on excitatory neurons of the cortex contain &alpha4 subunits, whereas on interneurons they contain &alpha1 subunits. This allows to pharmacologically differentiate &delta-GABAARs on specific neuron. Development of drugs optimized to selectively modify &delta-GABAA-mediated tonic inhibition of excitatory or inhibitory neurons may lead to critical clinical applications.

    Cover page: Plasticity of GABAergic Inhibition
    • Thesis
    • Peer Reviewed
    UCLA Electronic Theses and Dissertations (2017)

    Numerous efforts have been made to illuminate the mechanisms underlying neurological and psychiatric disorders, in order to develop treatments for these diseases. With the advantage of transgenic mouse models to recapitulate human pathological alterations and to support drug screening, promising discoveries have been made over the past century. In this dissertation, I am focusing on Alzheimer’s disease (AD), which is a neurodegenerative disorder with significant public health concern. Using one of the newest mouse models of AD, the AppNL-F mouse model, I have investigated synaptic, network, and cognitive alterations related to this disease, with techniques from in vitro and in vivo electrophysiology for examining abnormalities in the brain to behavior tests for assessing the cognitive functions of the animals. In the meantime, novel quantitative approaches have been developed and applied to objectively interpret the experimental data. In AppNL-F homozygous mice, we have found synaptic alterations consistent with deficits in parvalbumin-expressing interneurons (PV INs). Moreover, through long-term in vivo local field potential (LFP) recording, we have discovered perturbed network activities associated with disrupted sleep patterns, which happened during a critical period in the disease progression and may lead to further cognitive deficits in these animals.

    Besides well-known cognitive dysfunctions such as loss of memory and reasoning as well as language difficulties, there is a second group of non-cognitive symptoms including psychiatric disorders and behavioral disturbances, such as depression, hallucination, delusions, and agitations(Burns et al., 1990; Burns and Iliffe, 2009). Therefore, I have also investigated into the mouse forced-swim test (FST), which is widely used in the screening of antidepressants. By quantitative analyses on this test, we have discovered a new confounding factor, and developed novel methods to determine the important parameters in the mouse FST, which could be applied to mouse AD models to study depression and the effect of antidepressants in AD.

    In conclusion, the application of quantitative approaches in neuroscience studies have led to the discovery of intriguing results and provided ground for exciting future research.

      Cover page: Novel Quantitative Analyses in Mouse Models of Neurological and Psychiatric Disorders
      • Thesis
      • Peer Reviewed
      UCLA Electronic Theses and Dissertations (2014)

      Much work has shown that Parvalbumin+ (PV) interneurons, in particular the Fast Spiking Basket Cell, are major regulators of gamma-band activity in the cortex and hippocampus. Gamma rhythms, which are linked to attention, learning, and cognition, are depressed or altered in several key neurologic and psychiatric conditions, including schizophrenia and autism. Considerable evidence has demonstrated PV interneurons to be rigid regulators of neuronal activity, similar to a clock or a timer; however, in the past several years, evidence points to the ability of these cells to exert considerable modulatory influence on networks. For example, PV interneurons are surrounded by rings of dense extracellular matrix, termed perineuronal nets (PNN), which regulate plasticity in the visual cortex during development, as well as long-term potentiation and memory in the limbic lobe. Additionally, PV interneurons express extrasynaptic delta-subunit containing GABAA receptors (delta-GABAARs) that mediate tonic inhibition. In addition to regulating the excitability of these cells, delta-GABAARs are sensitive to levels of metabolites of progesterone and adrenal corticoids (i.e., neurosteroids), which are potent allosteric modulators of these receptors.

      Here, through a pharmacological and genetic model, we investigated the role of PV interneuron-expressed delta-GABAARs in behavior and neural rhythms. We show that ketamine, a non-competitive NMDA receptor antagonist shown to damage hippocampal PV interneurons, reduces and alters the PNNs surrounding these cells, as well as the expression of PV, and produces a behavioral phenotype consistent with schizophrenia. We next investigated the effect of conditionally knocking out the delta-GABAA receptor from PV interneurons. This resulted in altered gamma frequency and sensorimotor gating. Collectively, these results indicate that PV interneurons are capable of modulating cognition and behavior through tonic inhibitory currents. While other studies have demonstrated a role of tonic inhibition in behavior before, this is the first study to demonstrate interneuron-specific consequences of reduced delta-GABAAR expression.

        Cover page: Genetic and Pharmacological Manipulations of Parvalbumin Interneurons: Relevance to Neuropsychiatric Disease
        • Article
        • Peer Reviewed
        UCLA Previously Published Works (2010)
        Perisomatic inhibition from basket cells plays an important role in regulating pyramidal cell output. Two major subclasses of CA1 basket cells can be identified based on their expression of either cholecystokinin (CCK) or parvalbumin. This study examined their fates in the mouse pilocarpine model of temporal lobe epilepsy. Overall, immunohistochemical labeling of GABAergic boutons in the pyramidal cell layer of CA1 was preserved in the mouse model. However, CCK-labeled boutons in this layer were chronically reduced, whereas parvalbumin-containing boutons were conserved. Immunohistochemistry for cannabinoid receptor 1 (CB(1)), another marker for CCK-containing basket cells, also labeled fewer boutons in pilocarpine-treated mice. Hours after status epilepticus, electron microscopy revealed dark degenerating terminals in the pyramidal cell layer with lingering CCK and CB(1) immunoreactivity. In mice with recurrent seizures, carbachol-induced enhancement of spontaneous IPSCs (sIPSCs) originating from CCK-containing basket cells was accordingly reduced in CA1 pyramidal cells. By suppressing sIPSCs from CCK-expressing basket cells, a CB(1) agonist reverted the stimulatory effects of carbachol in naive mice to levels comparable with those observed in cells from epileptic mice. The agatoxin-sensitive component of CA1 pyramidal cell sIPSCs from parvalbumin-containing interneurons was increased in pilocarpine-treated mice, and miniature IPSCs were reduced, paralleling the decrease in CCK-labeled terminals. Altogether, the findings are consistent with selective reduction in perisomatic CA1 pyramidal cell innervation from CCK-expressing basket cells in mice with spontaneous seizures and a greater reliance on persisting parvalbumin innervation. This differential alteration in inhibition may contribute to the vulnerability of the network to seizure activity.
          Creative Commons 'BY' version 4.0 license
          • Thesis
          • Peer Reviewed
          UCLA Electronic Theses and Dissertations (2014)

          Temporal lobe epilepsy (TLE) is a form of acquired epilepsy characterized by recurrent and unprovoked seizures. TLE often develops following a precipitating neurological insult, such as traumatic brain injury, stroke, infection, prolonged febrile seizures or status epilepticus. These insults can initiate a constellation of genetic, functional, network and systems level reorganization that transforms a normal non-epileptic brain into one capable of generating recurrent and unprovoked seizures. The period of time between neurological insults and the onset of epilepsy is known as the latent period and is believed to be the period of time during which underlying epileptogenic processes occur.

          The dentate gyrus (DG) region of the hippocampus plays a key role in contextual learning and memory and strongly regulates the flow of neural activity from cortical regions into seizure-prone hippocampal regions through emergent properties of its neural microcircuit. In TLE, this filtering function is disrupted and can contribute to generation of seizures. Recent studies have described pathological high-frequency oscillations (pHFOs) in the DG and identified them as potential biomarkers of underlying proconvulsant network abnormalities. Moreover, pHFOs appear to be harbingers of TLE and are generated in the DG prior to the onset of an epileptic phenotype and therefore may reflect underlying network reorganization

          during the epileptogenesis. Currently, little is known about the emergence of pHFOs after epilepsy-inducing neuronal insults or the pathological microcircuit that generates them. To address this we used a new mouse model of TLE to track the temporal and spectral evolution of pHFOs during epileptogenesis. Moreover, we investigated how disrupted integration of newly generated granule cells into the DG during epileptogenesis contributes to the disruption of DG microcircuit function.

          We found that DG pHFOs are generated in bursts in vivo during epileptogenesis but that their burst properties do not evolve over time. In contrast, pHFO spectral dynamics appear to evolve during the latent period, suggesting that pHFO properties change according to the epileptogenic stage. In vitro electrophysiologcal and optogenetic experiments revealed that NGCs abnormally integrate into the DG microcircuit during epileptogenesis and that optogenetic activation of these neurons is sufficient to induce pHFOs in the DG.

            Cover page: Reorganization of Dentate Gyrus Microcircuits During Epileptogenesis
            • Article
            • Peer Reviewed
            UCLA Previously Published Works (2018)
            Potassium ions significantly contribute to the resting membrane potential of cells and, therefore, extracellular K+ concentration is a crucial regulator of cell excitability. Altered concentrations of extracellular K+ affect the resting membrane potential and cellular excitability by shifting the equilibria between closed, open and inactivated states for voltage-dependent ion channels that underlie action potential initiation and conduction. Hence, it is valuable to directly measure extracellular K+ dynamics in health and diseased states. Here, we describe how to make, calibrate and use monopolar K+-selective microelectrodes. We deployed them in adult hippocampal brain slices to measure electrically evoked K+ concentration dynamics. The judicious use of such electrodes is an important part of the tool-kit needed to evaluate cellular and biophysical mechanisms that control extracellular K+ concentrations in the nervous system.
              Cover page: Making, Testing, and Using Potassium Ion Selective Microelectrodes in Tissue Slices of Adult Brain.
              • Article
              • Peer Reviewed
              UCLA Previously Published Works (2021)
              Mossy cells (MCs) of the dentate gyrus (DG) are a major group of excitatory hilar neurons that are important for regulating activity of dentate granule cells. MCs are particularly intriguing because of their extensive longitudinal connections within the DG. It has generally been assumed that MCs in the dorsal and ventral DG have similar patterns of termination in the inner one-third of the dentate molecular layer. Here, we demonstrate that axonal projections of MCs in these two regions are considerably different. MCs in dorsal and ventral regions were labeled selectively with Cre-dependent eYFP or mCherry, using two transgenic mouse lines (including both sexes) that express Cre-recombinase in MCs. At four to six weeks following unilateral labeling of MCs in the ventral DG, a dense band of fibers was present in the inner one-fourth of the molecular layer and extended bilaterally throughout the rostral-caudal extent of the DG, replicating the expected distribution of MC axons. In contrast, following labeling of MCs in the dorsal DG, the projections were more diffusely distributed. At the level of transfection, fibers were present in the inner molecular layer, but they progressively expanded into the middle molecular layer and, most ventrally, formed a distinct band in this region. Optical stimulation of these caudal fibers expressing ChR2 demonstrated robust EPSCs in ipsilateral granule cells and enhanced the effects of perforant path stimulation in the ventral DG. These findings suggest that MCs in the dorsal and ventral DG differ in the distribution of their axonal projections and possibly their function.SIGNIFICANCE STATEMENT Mossy cells (MCs), a major cell type in the hilus of the dentate gyrus (DG), are unique in providing extensive longitudinal and commissural projections throughout the DG. Although it has been assumed that all MCs have similar patterns of termination in the inner molecular layer of the DG, we discovered that the axonal projections of dorsal and ventral MCs differ. While ventral MC projections exhibit the classical pattern, with dense innervation in the inner molecular layer, dorsal MCs have a more diffuse distribution and expand into the middle molecular layer where they overlap and interact with innervation from the perforant path. These distinct locations and patterns of axonal projections suggest that dorsal and ventral MCs may have different functional roles.
                Creative Commons 'BY' version 4.0 license
                • Article
                • Peer Reviewed
                UCLA Previously Published Works (2023)
                Subcortical white matter stroke (WMS) is a progressive disorder which is demarcated by the formation of small ischemic lesions along white matter tracts in the CNS. As lesions accumulate, patients begin to experience severe motor and cognitive decline. Despite its high rate of incidence in the human population, our understanding of the cause and outcome of WMS is extremely limited. As such, viable therapies for WMS remain to be seen. This study characterizes myelin recovery following stroke and motor learning-based rehabilitation in a mouse model of subcortical WMS. Following WMS, a transient increase in differentiating oligodendrocytes occurs within the peri-infarct in young male adult mice, which is completely abolished in male aged mice. Compound action potential recording demonstrates a decrease in conduction velocity of myelinated axons at the peri-infarct. Animals were then tested on one of three distinct motor learning-based rehabilitation strategies (skilled reach, restricted access to a complex running wheel, and unrestricted access to a complex running wheel) for their capacity to induce repair. These studies determined that unrestricted access to a complex running wheel alone increases the density of differentiating oligodendrocytes in infarcted white matter in young adult male mice, which is abolished in aged male mice. Unrestricted access to a complex running wheel was also able to enhance conduction velocity of myelinated axons at the peri-infarct to a speed comparable to naive controls suggesting functional recovery. However, there was no evidence of motor rehabilitation-induced remyelination or myelin protection.SIGNIFICANCE STATEMENT White matter stroke is a common disease with no medical therapy. A form of motor rehabilitation improves some aspects of white matter repair and recovery.
                  Cover page: Motor Activity-Induced White Matter Repair in White Matter Stroke.
                  • Article
                  • Peer Reviewed
                  UCLA Previously Published Works (2023)

                  Introduction

                  Various methods have been used to determine the frequency components of seizures in scalp electroencephalography (EEG) and in intracortical recordings. Most of these methods rely on subjective or trial-and-error criteria for choosing the appropriate bandwidth for filtering the EEG or local field potential (LFP) signals to establish the frequency components that contribute most to the initiation and maintenance of seizure activity. The empirical mode decomposition (EMD) with the Hilbert-Huang transform is an unbiased method to decompose a time and frequency variant signal into its component non-stationary frequencies. The resulting components, i.e., the intrinsic mode functions (IMFs) objectively reflect the various non-stationary frequencies making up the original signal.

                  Materials and methods

                  We employed the EMD method to analyze the frequency components and relative power of spontaneous electrographic seizures recorded in the dentate gyri of mice during the epileptogenic period. Epilepsy was induced in mice following status epilepticus induced by suprahippocampal injection of kainic acid. The seizures were recorded as local field potentials (LFP) with electrodes implanted in the dentate gyrus. We analyzed recording segments that included a seizure (mean duration 28 s) and an equivalent time period both before and after the seizure. Each segment was divided into non-overlapping 1 s long epochs which were then analyzed to obtain their IMFs (usually 8-10), the center frequencies of the respective IMF and their spectral root-mean-squared (RMS) power.

                  Results

                  Our analysis yielded unbiased identification of the spectral components of seizures, and the relative power of these components during this pathological brain activity. During seizures, the power of the mid frequency components increased while the center frequency of the first IMF (with the highest frequency) dramatically decreased, providing mechanistic insights into how local seizures are generated.

                  Discussion

                  We expect this type of analysis to provide further insights into the mechanisms of seizure generation and potentially better seizure detection.
                    Cover page: Capturing the power of seizures: an empirical mode decomposition analysis of epileptic activity in the mouse hippocampus.
                    • Article
                    • Peer Reviewed
                    UCLA Previously Published Works (2007)
                    Cooperativity is one of the most important properties of molecular interactions in biological systems. It is the ability to influence ligand binding at one site of a macromolecule by previous ligand binding at another site of the same molecule. As a consequence, the affinity of the macromolecule for the ligand is either decreased (negative cooperativity) or increased (positive cooperativity). Over the last 100 years, O2 binding to hemoglobin has served as the paradigm for cooperative ligand binding and allosteric modulation, and four practical models were developed to quantitatively describe the mechanism: the Hill, the Adair-Klotz, the Monod-Wyman-Changeux, and the Koshland-Némethy-Filmer models. The predictions of these models apply under static conditions when the binding reactions are at equilibrium. However, in a physiological setting, e.g., inside a cell, the timing and dynamics of the binding events are essential. Hence, it is necessary to determine the dynamic properties of cooperative binding to fully understand the physiological implications of cooperativity. To date, the Monod-Wyman-Changeux model was applied to determine the kinetics of cooperative binding to biologically active molecules. In this model, cooperativity is established by postulating two allosteric isoforms with different binding properties. However, these studies were limited to special cases, where transition rates between allosteric isoforms are much slower than the binding rates or where binding and unbinding rates could be measured independently. For all other cases, the complex mathematical description precludes straightforward interpretations. Here, we report on calculating for the first time the fast dynamics of a cooperative binding process, the binding of Ca2+ to calretinin. Calretinin is a Ca2+-binding protein with four cooperative binding sites and one independent binding site. The Ca2+ binding to calretinin was assessed by measuring the decay of free Ca2+ using a fast fluorescent Ca2+ indicator following rapid (<50-mus rise time) Ca2+ concentration jumps induced by uncaging Ca2+ from DM-nitrophen. To unravel the kinetics of cooperative binding, we devised several approaches based on known cooperative binding models, resulting in a novel and relatively simple model. This model revealed unexpected and highly specific nonlinear properties of cellular Ca2+ regulation by calretinin. The association rate of Ca2+ with calretinin speeds up as the free Ca2+ concentration increases from cytoplasmic resting conditions ( approximately 100 nM) to approximately 1 muM. As a consequence, the Ca2+ buffering speed of calretinin highly depends on the prevailing Ca2+ concentration prior to a perturbation. In addition to providing a novel mode of action of cellular Ca2+ buffering, our model extends the analysis of cooperativity beyond the static steady-state condition, providing a powerful tool for the investigation of the dynamics and functional significance of cooperative binding in general.
                      Cover page: Resolving the Fast Kinetics of Cooperative Binding: Ca2+ Buffering by Calretinin
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