Shobhana Sivaramakrishnan

IDENTIFICATION OF CELL TYPES IN BRAIN SLICES OF THE INFERIOR COLLICULUS

Different type neurons in the inferior colliculus may have different functions. Recent intracellular studies of the inferior colliculus suggest that intrinsic electrical properties contribute to discharge patterns, but the intrinsic discharge patterns have not been fully characterized in the central nucleus, the main part of the inferior colliculus. Whether different types of neurons are related to different discharge patterns is unclear. We have used intracellular and whole-cell patch clamp-recording techniques in a brain slice preparation to better characterize discharge patterns and cell types in the central nucleus. Several types of discharge pattern were found in the inferior colliculus in response to long pulses of intracellular depolarizations. Rebound and buildup-pauser discharges, together, comprise neurons with a sustained response and are the majority of the neurons in the inferior colliculus. Both of these types of discharge pattern could be adapting or regular. Onset discharges distinguished another group of neurons. Onset neurons can also entrain to higher frequency stimuli than sustained neurons. Discharge patterns are correlated with distinctive current-voltage relationships and with some aspects of dendritic morphology. However, the morphological data demonstrates that the discharge patterns do not correspond simply to disc-shaped (flat) or stellate (less-flat) categories. This is the first extensive analysis of electrophysiological properties of the central nucleus of the inferior colliculus in vitro. We suggest that there may be at least three functional classes of neurons and have implications for signal processing in the inferior colliculus.

GABAA Synapses Shape Neuronal Responses to Sound Intensity in the Inferior Colliculus

Neurons in the inferior colliculus (IC) change their firing rates with sound pressure level. Some neurons maintain monotonic increases in firing rate over a wide range of sound intensities, whereas other neurons are monotonic over limited intensity ranges. We examined the conditions necessary for monotonicity in this nucleus in vitro in rat brain slices and in vivo in the unanesthetized rabbit. Our in vitro recordings indicate that concurrent activation of GABAA synapses with excitatory inputs facilitates monotonic increases in firing rate with increases in stimulus strength. In the absence of synaptic inhibition, excitatory input to IC neurons causes large depolarizations that result in firing block and nonmonotonicity. In vivo, although GABAA synapses decrease the firing rate in all IC neurons, they can have opposing effects on rate-level functions. GABAergic inputs activated by all sound intensities maintain monotonicity by keeping the postsynaptic potential below the level at which depolarization block occurs. When these inputs are blocked, firing block can occur and rate-level functions become nonmonotonic. High-threshold GABAergic inputs, in contrast, cause nonmonotonic responses by decreasing the firing rate at high intensities. Our results suggest that a dynamic regulation of the postsynaptic membrane potential by synaptic inhibition is necessary to allow neurons to respond monotonically to a wide range of sound intensities.

Neuronal Responses to Lemniscal Stimulation in Laminar Brain Slices of the Inferior Colliculus

The central nucleus of the inferior colliculus (ICC) receives inputs from all parts of the auditory brainstem and transmits the information to the forebrain. Fibrodendritic laminae of the ICC provide a structural basis for a tonotopic organization, and the interaction of inputs within a single layer is important for ICC processing. Transverse slice planes of the ICC sever the layers and many of the ascending axons that enter through the lateral lemniscus. Consequently, the activity initiated within a lamina by a pure lemniscal stimulus is not well characterized. Here, we use a slice plane that maintains the integrity of the laminae in ICC and allows the axons in the lateral lemniscus to be stimulated at a distance from the ICC. We examined both the postsynaptic currents and potentials of the same neurons to lemniscal stimuli in this laminar brain slice. Our main finding is that lemniscal stimulation evokes prolonged synaptic potentials in ICC neurons. Synaptic potential amplitudes and durations increase with lemniscal shock strength. In ∼50% of ICC neurons, the postsynaptic potential is equal in duration to the postsynaptic current, whereas in the remaining neurons it is three to four times longer. Synaptic responses to single shocks or shock trains exhibit plateau potentials that enable sustained firing in ICC neurons. Plateau potentials are evoked by N-methyl-d-aspartate (NMDA) receptor activation, and their amplitudes and durations are regulated by both NMDA-R and gamma-aminobutyric acid A (GABAA)-R activation. These data suggest that in the intact laminae of the ICC, lemniscal inputs initiate sustained firing through monosynaptic and polysynaptic NMDA-mediated synapses regulated by GABAA synapses.

Haploinsufficiency in Peptidylglycine α-Amidating Monooxygenase Leads to Altered Synaptic Transmission in the Amygdala and Impaired Emotional Responses

The mammalian amygdala expresses various neuropeptides whose signaling has been implicated in emotionality. Many neuropeptides require amidation for full activation by peptidylglycine α-amidating monooxygenase (PAM), a transmembrane vesicular cuproenzyme and regulator of the secretory pathway. Mice heterozygous for the Pam gene (PAM+/−) exhibit physiological and behavioral abnormalities related to specific peptidergic pathways. In the present study, we evaluated emotionality and examined molecular and cellular responses that characterize neurophysiological differences in the PAM+/− amygdala. PAM+/− mice presented with anxiety-like behaviors in the zero maze that were alleviated by diazepam. PAM+/− animals were deficient in short- and long-term contextual and cued fear conditioning and required higher shock intensities to establish fear-potentiated startle than their wild-type littermates. Immunohistochemical analysis of the amygdala revealed PAM expression in pyramidal neurons and local interneurons that synthesize GABA. We performed whole-cell recordings of pyramidal neurons in the PAM+/− amygdala to elucidate neurophysiological correlates of the fear behavioral phenotypes. Consistent with these observations, thalamic afferent synapses in the PAM+/− lateral nucleus were deficient in long-term potentiation. This deficit was apparent in the absence and presence of the GABAA receptor antagonist picrotoxin and was abolished when both GABAA and GABAB receptors were blocked. Both evoked and spontaneous excitatory signals were enhanced in the PAM+/− lateral nucleus. Phasic GABAergic signaling was also augmented in the PAM+/− amygdala, and this difference comprised activity-independent and -dependent components. These physiological findings represent perturbations in the PAM+/− amygdala that may underlie the aberrant emotional responses in the intact animal.

Functional architecture of the inferior colliculus revealed with voltage-sensitive dyes.

We used optical imaging with voltage-sensitive dyes to investigate the spatio-temporal dynamics of synaptically evoked activity in brain slices of the inferior colliculus (IC). Responses in transverse slices which preserve cross-frequency connections and in modified sagittal slices that preserve connections within frequency laminae were evoked by activating the lateral lemniscal tract. Comparing activity between small and large populations of cells revealed response areas in the central nucleus of the IC that were similar in magnitude but graded temporally. In transverse sections, these response areas are summed to generate a topographic response profile. Activity through the commissure to the contralateral IC required an excitation threshold that was reached when GABAergic inhibition was blocked. Within laminae, module interaction created temporal homeostasis. Diffuse activity evoked by a single lemniscal shock re-organized into distinct spatial and temporal compartments when stimulus trains were used, and generated a directional activity profile within the lamina. Using different stimulus patterns to activate subsets of microcircuits in the central nucleus of the IC, we found that localized responses evoked by low-frequency stimulus trains spread extensively when train frequency was increased, suggesting recruitment of silent microcircuits. Long stimulus trains activated a circuit specific to post-inhibitory rebound neurons. Rebound microcircuits were defined by a focal point of initiation that spread to an annular ring that oscillated between inhibition and excitation. We propose that much of the computing power of the IC is derived from local circuits, some of which are cell-type specific. These circuits organize activity within and across frequency laminae, and are critical in determining the stimulus-selectivity of auditory coding.

Midbrain local circuits shape sound intensity codes

Hierarchical processing of sensory information requires interaction at multiple levels along the peripheral to central pathway. Recent evidence suggests that interaction between driving and modulating components can shape both top down and bottom up processing of sensory information. Here we show that a component inherited from extrinsic sources combines with local components to code sound intensity. By applying high concentrations of divalent cations to neurons in the nucleus of the inferior colliculus in the auditory midbrain, we show that as sound intensity increases, the source of synaptic efficacy changes from inherited inputs to local circuits. In neurons with a wide dynamic range response to intensity, inherited inputs increase firing rates at low sound intensities but saturate at mid-to-high intensities. Local circuits activate at high sound intensities and widen dynamic range by continuously increasing their output gain with intensity. Inherited inputs are necessary and sufficient to evoke tuned responses, however local circuits change peak output. Push–pull driving inhibition and excitation create net excitatory drive to intensity-variant neurons and tune neurons to intensity. Our results reveal that dynamic range and tuning re-emerge in the auditory midbrain through local circuits that are themselves variable or tuned.

High concentrations of divalent cations isolate monosynaptic inputs from local circuits in the auditory midbrain.

Hierarchical processing of sensory information occurs at multiple levels between the peripheral and central pathway. Different extents of convergence and divergence in top down and bottom up projections makes it difficult to separate the various components activated by a sensory input. In particular, hierarchical processing at sub-cortical levels is little understood. Here we have developed a method to isolate extrinsic inputs to the inferior colliculus (IC), a nucleus in the midbrain region of the auditory system, with extensive ascending and descending convergence. By applying a high concentration of divalent cations (HiDi) locally within the IC, we isolate a HiDi-sensitive from a HiDi-insensitive component of responses evoked by afferent input in brain slices and in vivo during a sound stimulus. Our results suggest that the HiDi-sensitive component is a monosynaptic input to the IC, while the HiDi-insensitive component is a local polysynaptic circuit. Monosynaptic inputs have short latencies, rapid rise times, and underlie first spike latencies. Local inputs have variable delays and evoke long-lasting excitation. In vivo, local circuits have variable onset times and temporal profiles. Our results suggest that high concentrations of divalent cations should prove to be a widely useful method of isolating extrinsic monosynaptic inputs from local circuits in vivo.

Postnatal developmental changes in the medial nucleus of the trapezoid body in a mouse model of auditory pathology

Age-related hearing loss (AHL) is a multifactorial disorder characterized by a decline in peripheral and central auditory function. Here, we examined synaptic transmission in DBA/2 mice, which carry the AHL8 gene, at the identifiable glutamatergic synapse in the medial nucleus of the trapezoid body (MNTB), a nucleus in the superior olivary complex critical for acoustic timing. Mice exhibited raised auditory brainstem thresholds by P14, soon after hearing onset. Excitatory postsynaptic currents were prolonged; however, postsynaptic excitability was normal. By P18, high-frequency hearing loss was evident. Coincident with the onset of hearing loss, MNTB principal neurons displayed changes in intrinsic firing properties. These results suggest that changes in transmission in the superior olivary complex are associated with early onset hearing loss.

Postinhibitory rebound neurons and networks are disrupted in retrovirus-induced spongiform neurodegeneration

Certain retroviruses induce progressive spongiform motor neuron disease with features resembling prion diseases and amyotrophic lateral sclerosis. With the neurovirulent murine leukemia virus (MLV) FrCasE, Env protein expression within glia leads to postsynaptic vacuolation, cellular effacement, and neuronal loss in the absence of neuroinflammation. To understand the physiological changes associated with MLV-induced spongiosis, and its neuronal specificity, we employed patch-clamp recordings and voltage-sensitive dye imaging in brain slices of the mouse inferior colliculus (IC), a midbrain nucleus that undergoes extensive spongiosis. IC neurons characterized by postinhibitory rebound firing (PIR) were selectively affected in FrCasE-infected mice. Coincident with Env expression in microglia and in glia characterized by NG2 proteoglycan expression (NG2 cells), rebound neurons (RNs) lost PIR, became hyperexcitable, and were reduced in number. PIR loss and hyperexcitability were reversed by raising internal calcium buffer concentrations in RNs. PIR-initiated rhythmic circuits were disrupted, and spontaneous synchronized bursting and prolonged depolarizations were widespread. Other IC neuron cell types and circuits within the same degenerative environment were unaffected. Antagonists of NMDA and/or AMPA receptors reduced burst firing in the IC but did not affect prolonged depolarizations. Antagonists of L-type calcium channels abolished both bursts and slow depolarizations. IC infection by the nonneurovirulent isogenic virus Friend 57E (Fr57E), whose Env protein is structurally similar to FrCasE, showed no RN hyperactivity or cell loss; however, PIR latency increased. These findings suggest that spongiform neurodegeneration arises from the unique excitability of RNs, their local regulation by glia, and the disruption of this relationship by glial expression of abnormal protein.

Rebound from Inhibition: Self-Correction against Neurodegeneration?

Neural networks play a critical role in establishing constraints on excitability in the central nervous system. Several recent studies have suggested that network dysfunction in the brain and spinal cord are compromised following insult by a neurodegenerative trigger and might precede eventual neuronal loss and neurological impairment. Early intervention of network excitability and plasticity might therefore be critical in resetting hyperexcitability and preventing later neuronal damage. Here, the behavior of neurons that generate burst firing upon recovery from inhibitory input or intrinsic membrane hyperpolarization (rebound neurons) is examined in the context of neural networks that underlie rhythmic activity observed in areas of the brain and spinal cord that are vulnerable to neurodegeneration. In a non-inflammatory rodent model of spongiform neurodegenerative disease triggered by retrovirus infection of glia, rebound neurons are particularly vulnerable to neurodegeneration, likely due to an inherently low calcium buffering capacity. The dysfunction of rebound neurons translates into a dysfunction of rhythmic neural circuits, compromising normal neurological function and leading to eventual morbidity. Understanding how virus infection of glia can mediate dysfunction of rebound neurons, induce hyperexcitability and loss of rhythmic function, pathologic features observed in neurodegenerative disorders ranging from epilepsy to motor neuron disease, might therefore suggest a common pathway for early therapeutic intervention.