Tonic And Phasic Inhibitory Mechanisms Mediating Sensorimotor Decision-Making In The Goldfish Auditory Startle Circuit

This work describes related studies of cellular and synaptic signaling mechanisms involved in the balance of excitation and inhibition in the goldfish auditory startle circuit. The general purpose of these experiments was to identify novel mechanisms that contribute to action selection at different stages of the motor control hierarchy. The methods applied to achieve this goal tested the effects of selective antagonists for target receptor systems on sound-evoked excitation and inhibition of startle. Chapter 2 describes a study of a poorly-understood serotonergic mechanism, the 5-HT5A receptor, that was not previously functionally characterized in native tissues or associated with neural or behavioral processes. Treatment with a selective 5-HT5A antagonist caused a 26.41 ± 3.98% reduction in sound-evoked excitation of startle. Subsequent experiments revealed that the 5-HT5A antagonist significantly reduced post-synaptic excitability in the Mauthner-cell (M-cell) neurons that initiate startle. Despite these effects, prepulse inhibition (PPI) of the startle response remained robustly intact after treatment with the 5-HT5A antagonist. The 5-HT5A receptor is thus not a likely mechanism for PPI, but does act as a selective modulator of startle excitability. A final series of experiments confirmed that the 5-HT5A antagonist reduced M-cell excitability by increasing Cl- conductance, likely by activating Cl- channels. Chapter 3 presents experiments focused on the inhibitory neurotransmitters that directly mediate the phasic inhibitory process elicited during PPI. Strychnine, a glycine receptor (GlyR) antagonist, caused an 87.43 ± 21.53% increase in sound-evoked excitation of startle, but PPI remained robustly intact, despite this. GlyRs thus likely mediate a tonic inhibitory process that was blocked by strychnine treatment, but glycinergic components of sound-evoked inhibition decayed too rapidly (\u3c50 \u3ems) to contribute to the prolonged time-course of PPI. In a parallel series of experiments, treatment with bicuculline, the GABAAR antagonist, caused similar increases in sound-evoked excitation (by 133.8 ± 10.3%) of startle, but the GABAAR antagonist also significantly reduced auditory PPI at inter-stimulus intervals of 100 ms and less. In sum, these findings indicate that glycine and GABA tonically inhibit the M-cell startle circuit, but GABA is also the primary effector mechanism for inhibitory signaling during PPI. In summary, three goals were accomplished. First, the thorough functional characterization of 5-HT5A provides a fully integrated serotonergic mechanism, and this appears to provide an ideal tool for selective potentiation of startle. Next, experiments with strychnine emphasize a short-lived role of GlyRs in sound-evoked (feed-forward) inhibition, and also act as mediators of a tonic inhibitory process that controls startle excitability. Last, experiments with bicuculline identify GABA as the inhibitory neurotransmitter that directly mediates PPI

Noise-induced cochlear neuronal degeneration and its role in hyperacusis -- and tinnitus-like behavior

Thesis (Ph. D. in Speech and Hearing Bioscience and Technology)--Harvard-MIT Program in Health Sciences and Technology, 2013.Cataloged from PDF version of thesis.Includes bibliographical references (p. 46-57).Perceptual abnormalities such as hyperacusis and tinnitus often occur following acoustic overexposure. Although such exposure can also result in permanent threshold elevation, some individuals with noise-induced hyperacusis or tinnitus show clinically normal thresholds. Recent work in animals has shown that noise exposure can cause permanent degeneration of the cochlear nerve despite complete threshold recovery and lack of hair cell damage (Kujawa and Liberman, J Neurosci 29:14077-14085, 2009). Here we ask whether this noise-induced primary neuronal degeneration results in abnormal auditory behavior, indexed by the acoustic startle response and prepulse inhibition (PPI) of startle. Responses to tones and to broadband noise were measured in mice exposed either to a neuropathic exposure causing primary neuronal degeneration, or to a lower intensity, nonneuropathic noise, and in unexposed controls. Mice with cochlear neuronal loss displayed hyper-responsivity to sound, as evidenced by lower startle thresholds and enhanced PPI, while exposed mice without neuronal loss showed control-like responses. Gap PPI tests, often used to assess tinnitus, revealed spectrally restricted, as well as broadband, gap-detection deficits in mice with primary neuronal degeneration, but not in exposed mice without neuropathy. Crossmodal PPI tests and behavioral assays of anxiety revealed no significant differences among groups, suggesting that the changes in startle-based auditory behavior reflect a neuropathyrelated alteration specifically of auditory neural pathways. Despite significantly reduced cochlear nerve response, seen as reduced wave 1 of the auditory brainstem response, later peaks were unchanged or enhanced, suggesting neural hyperactivity in the auditory brainstem that could underlie the abnormal behavior on the startle tests. Taken together, the results suggest a role for cochlear primary neuronal degeneration in central neural excitability and, by extension, in the generation of tinnitus and hyperacusis.by Ann E. Hickox.Ph.D.in Speech and Hearing Bioscience and Technolog

The startle response as a measure in mouse models of mood disorders

Ein großer Teil der Fragestellungen in den Neurowissenschaften beschäftigt sich mit dem Thema, wie das Säugerhirn Verhalten auslöst und steuert. Die Schreckreaktion ist ein relativ einfaches Verhalten, welches bei Säugetieren ohne großen Aufwand ausgelöst werden kann und variabel auf eine Vielfalt von experimentellen Behandlungen reagiert. Das Ziel der vorliegenden Arbeit war es, Schreckreaktions-Messungen am Max-Planck- Institut für Psychiatrie in München (MPI-P) zu etablieren. Vor dem Hintergrund aktueller Fragestellungen sollten die Experimente zu einsatzbereiten Messmethoden und Verhaltensparadigmen führen. In der vorliegenden Arbeit gelang es nicht, das Paradigma der furchtpotenzierten Schreckreaktion (FPS) zuverlässig in einem häufig am MPI-P eingesetzten Mausstamm anzuwenden. Das FPS maskierende Phänomen, daß die Präsentation eines unkonditionierten Tons bereits zu einer deutlich verstärkten Schreckreaktion in diesen Mäusen führt ("tone enhanced startle", TES) wurde dann charakterisiert und im Folgenden als ergänzendes Paradigma zur Messung und Abschätzung des Hörvermögens, der Stimulus Adaptation und der Aufmerksamkeit in Mäusen vorgeschlagen. Eine Literaturrecherche ergab, daß im Paradigma der Furchtkonditionierung ("fear conditioning", FC) und deren aktives Verlernen ("extinction of FC", ExFC) verwendete Stimulus-Parameter eine hohe Varianz zwischen verschiedenen Laboratorien aufweisen. Der im Verhalten ausgelesene Lernerfolg während einer FC wie auch einem ExFC hingen in den vorliegenden Experimenten wesentlich von der verwendeten Stimulusqualität ab (d.h. sinus-Ton oder weißes Rauschen). Im Umkehrschluß empfiehlt die vorliegende Arbeit einen überlegteren Umgang mit den eingetzten Stimulus-Parametern. Es zeigte sich, daß eine erhöhte Schreckreaktion (Übererregbarkeit) ohne weiteres in einem Tiermodell der Posttraumatischen Belastungsstörung ("posttraumatic stress disorder",PTSD) gemessen werden kann. Im Weiteren konnte gezeigt werden, daß verändertes Hippocampus-Volumen in diesen Tieren, gemessen über ultramikroskopische Aufnahmen und analog zu Hippocampusveränderungen in Patienten, unabhängig von anderen PTSD-ähnlichen Symptomen dieser Mäuse ist. In einem weiteren Abschnitt widmet sich die vorliegende Arbeit der laufenden Charakterisierung der Rolle von Dopaminrezeptoren (DR) in der Präpulsinhibition (PPI) und -Faszilitierung (PPF) der Schreckreaktion. Durch lokale injektion von DR-Antagonisten konnte gezeigt werden, daß die Blockade von DR1 wiederholbar PPI verstärkt, während die Rolle von DR2, getestet mit zwei verschiedenen Antagonisten, als ambivalent gedeutet werden muß. Basierend auf diesen Experimenten wurden optogenetische Methoden in die Schreckreaktionsmessung eingeführt. Transgenen Mäusen, die lichtsensitive Ionenkanäle in ihren neuronalen Zellmembranen bestimmter Zellpopulationen tragen, wurden Lichtblitze ins Gehirn appliziert. Auf diese Weise konnten PPI und PPF unabhängig voneinander manipuliert werden. Daraus folgend, und im Unterschied zur populären Summationshypothese der PPF, schlägt die vorliegende Arbeit einen eigenständigen, von der PPI unabhängigen PPF-Schaltkreis vor, der Pyramidenneuronen der präfrontalen Kortexschicht V beinhaltet. Die vorliegende Arbeit konnte erfolgreich verschiedene Protokolle und Verhaltensparadigmen der Schreckreaktionsmessung am MPI-P etablieren und zur sofortigen Nutzung zur Verfügung stellen. Es wurden nicht nur neue Techniken wie z.B. optogenetische Methoden in die Schreckreaktionsmessung eingeführt, die vorliegenden Experiemente leisten auch ihren Beitrag zur aktiven Forschung, in dem sie z.B. die große Bedeutung der Stimulus-Parameter für den Lernerfolg von Versuchstieren nachweisen.In neuroscience major efforts are focused on the question of how the mammalian brain generates and controls behaviour. The startle response is a relatively simple behaviour that can be easily elicited in mammals and is sensitive to a variety of experimental treatments. The aim of the present work was to establish startle response measures at the Max-Planck-Institute of Psychiatry (MPI-P), Munich, providing a set of readily applicable methods and paradigms, and contributing to questions in behavioural neuroscience. While the present thesis failed to robustly elicit fear potentiated startle (FPS) in a commonly used mouse strain at the MPI-P, strong unconditioned startle enhancement by acoustic stimulus presentation in that mouse strain was capitalised to propose tone enhanced startle (TES) as an additional paradigm to assess hearing capability, stimulus adaptation and attention in mice. A literature survey revealed considerably varying parameters used in fear conditioning (FC) and extinction of conditioned fear (ExFC). In the present work, FC, ExFC as well as FPS/TES highly depended on the stimulus quality (i.e. sine wave or white noise), demanding a more careful handling of stimulus parameters. Hyper-arousal was readily tested in a mouse model of posttraumatic stress disorder (PTSD). Additionally it was shown that altered hippocampal volume in these animals, assessed by ultramicroscopic measures and mimicking patient data, was independent of other symptoms present in this model. The present thesis contributes to the ongoing characterisation of the role of dopamine receptors (DR) in prepulse inhibition (PPI) and prepulse facilitation (PPF) of startle, manipulating PPI/F by injections of DR-antagonists into the prefrontal cortex of mice. It was found that blockade of DR1 reliably increases PPI, while the effect of DR2 was inconsistent, using to different DR2-antagonists. Based on this work, optogenetic methods were established. Applying intracerebral light flashes to transgenic mice carrying light sensitive ion channels on their neuronal cell membrane, PPI and PPF were manipulated independently, proposing the existence of a discrete PPF mediating pathway including prefrontal layer V pyramidal neurons, contrasting the popular summation hypothesis of PPF. The present work established and developed successfully different startle paradigms that are ready to use for animal characterisation and testing. Apart from combining startle measures with new techniques such as optogenetic methods, the present thesis points out the stimulus parameter dependence of animal learning, suggesting a fundamental discussion about fear conditioning and extinction learning protocols

Acute stress impairs sensorimotor gating via the neurosteroid allopregnanolone in the prefrontal cortex

Ample evidence indicates that environmental stress impairs information processing, yet the underlying mechanisms remain partially elusive. We showed that, in several rodent models of psychopathology, the neurosteroid allopregnanolone (AP) reduces the prepulse inhibition (PPI) of the startle, a well-validated index of sensorimotor gating. Since this GABAA receptor activator is synthesized in response to acute stress, we hypothesized its participation in stress-induced PPI deficits. Systemic AP administration reduced PPI in C57BL/6J mice and Long-Evans, but not Sprague-Dawley rats. These effects were reversed by isoallopregnanolone (isoAP), an endogenous AP antagonist, and the GABAA receptor antagonist bicuculline and mimicked by AP infusions in the medial prefrontal cortex (mPFC). Building on these findings, we tested AP's implication in the PPI deficits produced by several complementary regimens of acute and short-term stress (footshock, restraint, predator exposure, and sleep deprivation). PPI was reduced by acute footshock, sleep deprivation as well as the combination of restraint and predator exposure in a time- and intensity-dependent fashion. Acute stress increased AP concentrations in the mPFC, and its detrimental effects on PPI were countered by systemic and intra-mPFC administration of isoAP. These results collectively indicate that acute stress impairs PPI by increasing AP content in the mPFC. The confirmation of these mechanisms across distinct animal models and several acute stressors strongly supports the translational value of these findings and warrants future research on the role of AP in information processing

Change in Acoustic Startle as an Indicator of Continuous Tonal Tinnitus

Currently, there is no accepted objective measure of tinnitus in humans. The gap prepulse inhibition of acoustic startle (GPIAS) paradigm is an objective measure that has been used in the animal model to identify tinnitus based on the theory of tinnitus filling in the silent gap that would normally promote startle inhibition. The current study applied the GPIAS paradigm in human subjects with normal hearing thresholds without hyperacusis. Individuals with continuous tonal tinnitus (N=31) characterized their tinnitus by adjusting a signal to match the frequency, bandwidth, and intensity. These individual parameters were used to create maximally matched background sounds in the GPIAS paradigm for each subject. A group without tinnitus (N=8) also participated using the averaged parameter values of the background sound from the group with tinnitus. Startle inhibition percentage was calculated by comparing ocular EMG blinking amplitudes between gap embedded conditions and the condition without a gap. As expected, the group with no tinnitus revealed startle inhibition as evidenced by reduced EMG blink amplitudes when the background sound was interrupted by a silent gap prior to the startle impulse (100 dB SPL white noise). The group with tinnitus did not have a significant startle inhibition in this same condition supporting the theory that the background sound carefully matched to their tinnitus eliminated the perception of a silent gap, thereby removing the cue that would produce startle inhibition. Gradually increasing the contrast between the individual’s continuous tonal tinnitus and ongoing background sound leads to a nonlinear change in startle inhibition percentage, providing guidelines for how closely the background sound needs to match the tinnitus of an individual in order to get the expected result of no startle inhibition when tinnitus is filling in the gap. Collectively, these findings support the use of the GPIAS paradigm for objectively identifying continuous tonal tinnitus in humans. Further, certain deviations in frequency, intensity, or bandwidth in the ongoing background sound from the tinnitus match result in startle inhibition, which may help explain the inconsistent findings across human GPIAS studies and allow more confidence for animal researchers to use GPIAS for animal tinnitus studies

The startle response as a measure in mouse models of mood disorders

In neuroscience major efforts are focused on the question of how the mammalian brain generates and controls behaviour. The startle response is a relatively simple behaviour that can be easily elicited in mammals and is sensitive to a variety of experimental treatments. The aim of the present work was to establish startle response measures at the Max-Planck-Institute of Psychiatry (MPI-P), Munich, providing a set of readily applicable methods and paradigms, and contributing to questions in behavioural neuroscience. While the present thesis failed to robustly elicit fear potentiated startle (FPS) in a commonly used mouse strain at the MPI-P, strong unconditioned startle enhancement by acoustic stimulus presentation in that mouse strain was capitalised to propose tone enhanced startle (TES) as an additional paradigm to assess hearing capability, stimulus adaptation and attention in mice. A literature survey revealed considerably varying parameters used in fear conditioning (FC) and extinction of conditioned fear (ExFC). In the present work, FC, ExFC as well as FPS/TES highly depended on the stimulus quality (i.e. sine wave or white noise), demanding a more careful handling of stimulus parameters. Hyper-arousal was readily tested in a mouse model of posttraumatic stress disorder (PTSD). Additionally it was shown that altered hippocampal volume in these animals, assessed by ultramicroscopic measures and mimicking patient data, was independent of other symptoms present in this model. The present thesis contributes to the ongoing characterisation of the role of dopamine receptors (DR) in prepulse inhibition (PPI) and prepulse facilitation (PPF) of startle, manipulating PPI/F by injections of DR-antagonists into the prefrontal cortex of mice. It was found that blockade of DR1 reliably increases PPI, while the effect of DR2 was inconsistent, using to different DR2-antagonists. Based on this work, optogenetic methods were established. Applying intracerebral light flashes to transgenic mice carrying light sensitive ion channels on their neuronal cell membrane, PPI and PPF were manipulated independently, proposing the existence of a discrete PPF mediating pathway including prefrontal layer V pyramidal neurons, contrasting the popular summation hypothesis of PPF. The present work established and developed successfully different startle paradigms that are ready to use for animal characterisation and testing. Apart from combining startle measures with new techniques such as optogenetic methods, the present thesis points out the stimulus parameter dependence of animal learning, suggesting a fundamental discussion about fear conditioning and extinction learning protocols

Central nervous system physiology

This is the second chapter of the series on the use of clinical neurophysiology for the study of movement disorders. It focusses on methods that can be used to probe neural circuits in brain and spinal cord. These include use of spinal and supraspinal reflexes to probe the integrity of transmission in specific pathways; transcranial methods of brain stimulation such as transcranial magnetic stimulation and transcranial direct current stimulation, which activate or modulate (respectively) the activity of populations of central neurones; EEG methods, both in conjunction with brain stimulation or with behavioural measures that record the activity of populations of central neurones; and pure behavioural measures that allow us to build conceptual models of motor control. The methods are discussed mainly in relation to work on healthy individuals. Later chapters will focus specifically on changes caused by pathology

Prepulse inhibition of the acoustic startle reflex: Comparing low and high trait anxious individuals

This investigation examined whether prepulse inhibition of the acoustic startle reflex was impaired among individuals with high trait anxiety compared to controls. PPI has traditionally been theorized to be a psychophysiological index of information processing and sensorimotor gating, and is now being used to help identify various psychological disorders. Although results of existing research on reduced PPI across the anxiety spectrum are equivocal, previous findings indicate that highly anxious (HA) participants exhibit significantly reduced PPI at lead intervals of 60 ms relative to low anxious (LA) controls. This study paired a 70 dB (A) white noise prepulse stimulus with a 100 dB (A) white noise startling stimulus at a 60 msec discrete lead interval. The results showed a higher baseline startle response and a trend toward increased response probability among the HA subgroup compared to controls. Due to a large number of nonresponse trials, analysis of PPI between groups could not be performed, but the data that was obtained revealed a 2.2% reduction in PPI among highly anxious participants. The current results highlight the potential for PPI to index sensorimotor gating deficits among anxious populations

Clerodendrum inerme

Previously, we found a patient with intractable motor tic disorder, a spectrum of Tourette syndrome (TS), responsive to the ground leaf juice of Clerodendrum inerme (CI). Here, we examined the effect of the ethanol extract of CI leaves (CI extract) on animal behaviors mimicking TS, hyperlocomotion, and sensorimotor gating deficit. The latter is also observed in schizophrenic patients and can be reflected by a disruption of prepulse inhibition of acoustic startle response (PPI) in animal models induced by methamphetamine and NMDA channel blockers (ketamine or MK-801), based on hyperdopaminergic and hypoglutamatergic hypotheses, respectively. CI extract (10–300 mg/kg, i.p.) dose-dependently inhibited hyperlocomotion induced by methamphetamine (2 mg/kg, i.p.) and PPI disruptions induced by methamphetamine, ketamine (30 mg/kg, i.p.), and MK-801 (0.3 mg/kg, i.p.) but did not affect spontaneous locomotor activity, rotarod performance, and grip force. These results suggest that CI extract can relieve hyperlocomotion and improve sensorimotor gating deficit, supporting the therapeutic potential of CI for TS and schizophrenia