Noise-induced hearing loss (NIHL) is a major cause of auditory processing impairment and major cognitive impairment. Peripherally, cochlear hair cell damage has been studied extensively in NIHL. Fewer studies have looked at effects more centrally, but these studies have led to the emerging theory of ‘central gain’. The lack of peripheral input (deafferentation) following NIHL increases gain in central auditory regions, leading to exaggerated spontaneous and sound evoked responses. These responses may lead to tinnitus and hyperacusis, for which currently there are no treatment to reduce these comorbidities following NIHL. The cellular mechanisms underlying the increased gain is unclear, however we hypothesize that neuroinflammation and matrix-metalloproteinase-9 (MMP-9) are upregulated following NIHL causing disruption in population level activity ultimately leading to enhanced central gain and temporal processing. To address this gap, we designed a series of experiments to identify the effects of NIHL in the brain. We have made several key findings. Firstly, there is a cortical-region specific difference in temporal processing, but not in sound detection and recovery following NIHL. Second, MMP-9 activity is not upregulated at 1-day post-NIHL. Last, minocycline enhances sound detection amplification after temporary hearing loss, but does not affect temporal processing or auditory brainstem thresholds. Our data provides insight into longitudinal modulations of sound detection and temporal processing in auditory cortical regions with potential timecourse and therapeutic targets.

The pallid bat (Antrozous pallidus), a gleaning bat found in the western United States and Mexico, hunts a wide variety of ground-dwelling prey, including scorpions. Anecdotal evidence suggests that the pallid bat is resistant to scorpion venom, but no systematic study has been performed. Here we show with behavioral measures and direct injection of venom that the pallid bat is resistant to venom of the Arizona bark scorpion, Centruroides sculpturatus. Our results show that the pallid bat is stung multiple times during a hunt without any noticeable effect on behavior. In addition, direct injection of venom at mouse LD50 concentrations (1.5 mg/kg) has no effect on bat behavior. At the highest concentration tested (10 mg/kg), three out of four bats showed no effects. One of the four bats showed a transient effect suggesting that additional studies are required to identify potential regional variation in venom tolerance. Scorpion venom is a cocktail of toxins, some of which activate voltage-gated sodium ion channels, causing intense pain. Dorsal root ganglia (DRG) contain nociceptive neurons and are principal targets of scorpion venom toxins. To understand if mutations in specific ion channels contribute to venom resistance, a pallid bat DRG transcriptome was generated. As sodium channels are a major target of scorpion venom, we identified amino acid substitutions present in the pallid bat that may lead to venom resistance. Some of these substitutions are similar to corresponding amino acids in sodium channel isoforms responsible for reduced venom binding activity. The substitution found previously in the grasshopper mouse providing venom resistance to the bark scorpion is not present in the pallid bat, indicating a potentially novel mechanism for venom resistance in the bat that remains to be identified. Taken together, these results indicate that the pallid bat is resistant to venom of the bark scorpion and altered sodium ion channel function may partly underlie such resistance.The pallid bat (Antrozous pallidus) is a gleaning bat found in western North America and is a well known hunter of scorpions. Previous work has shown the pallid bat is highly resistant to the venom of the Arizona barks scorpion, Centruroides sculpturatus. Here we build upon that work and show how the pallid bat may overcome the venom via cellular and genetic adaptations. Pallid bats were injected with doses of C. sculpturatus venom up to 20 mg/kg and showed only minor symptoms and made full recoveries. The serum neutralizing effects of pallid bat blood serum was tested via incubation of pallid bat blood serum with the venom before injection into mice. The serum incubated venom retained equivalent potency as the non-serum-incubated venom suggesting the pallid bat does not inactivate the venom via serum-based mechanisms. The sensory neurons in pallid bat dorsal root and trigeminal ganglia were tested via the Constellation Pharmacology method. Pallid bat sensory neurons experienced more calcium signal when challenged with C. sculpturatus venom than mouse sensory neurons, suggesting pallid bat venom resistance is not dependent on preventing increases in somatic calcium concentrations. The sequences of two voltage gated sodium channels were compared to other species and possible sites influencing venom action showed signs of positive selection in scorpion hunting bats.

Adaptations that allow for greater discrimination of low intensity sounds may be important in the echolocation behavior of bats. In this dissertation, we used the pallid bat, Antrozous pallidus, a gleaning bat of the western United States, as a model to study the adaptations for intensity selectivity present in the auditory cortex. We performed in-vivo extracellular recordings in the auditory cortex of the pallid bat, to determine the cortical organization and mechanisms of intensity selectivity. Downward frequency modulated (DFM) sweeps that approximated echolocation calls were used to study intensity selectivity using a behaviorally relevant sound. We also examined the distribution of Parvalbumin (PV) and Calbindin (CB) expressing cells in cortical regions and thalamic nuclei in the auditory pathway. Immunohistochemical staining was used to determine if the distribution of these calcium binding proteins in cortical and thalamic regions that have been implicated in processing of echolocation calls and how that distribution is different from non-echolocating regions. Lastly we examined the thalamocortical projections to intensity selective neurons in the echolocation region of the auditory cortex. Retrograde tracing from intensity selective and non-selective neurons was used to identify the thalamic nuclei that provided input to those neurons. We show that the region of the auditory cortex that is selective for echolocation calls contains a majority of neurons that are highly selective for low intensity sounds. We discovered that in the pallid bat intensity selectivity is enhanced by using behaviorally relevant stimuli and that high-frequency inhibition in the bat's echolocation call is responsible for this increased selectivity. This suggested a spectrotemporal integration mechanism that can shape intensity selectivity. Cortical mapping, in this study, revealed a systematic organization of intensity selectivity measures. We also discovered a differential staining pattern of calcium binding protein in the cortex as higher percentage of PV+ cells compared to CB+ cells was found in both echolocation call- and non-call regions. CB+ neurons where found in all of the regions of the medial geniculate body of the auditory thalamus, while PV staining was limited to the suprageniculate (SG), a region known to project to echolocation selective regions of the cortex. This study also confirmed previous results showing that echolocation selective regions of the cortex receive projections from all nuclei of the MGB except the lemniscal ventral nucleus, however an organization related to intensity selectivity was not able to be determined in the projections. The results from this dissertation highlight the special adaptations that are present in the auditory cortex of the pallid bat that may be important for processing the low-intensity echolocation call. Deviations from a general mammalian plan or even from other bat species in the properties that lead to these adaptations may strengthen the notion that general patterns of cortical processing and organization may be altered through evolution to support the unique behavioral needs of a species.

The population over the age of 65 years old is rapidly increasing around the world. Presbycusis (age-related hearing loss) is one of the most prevalent disabilities associated with age and can impair speech understanding and social and emotional well-being. Deficits in auditory processing, including speech understanding are likely caused by an interaction between changes in the way the brain processes sound and peripheral sensory hearing loss. It is difficult to disambiguate changes that occur within the brain independent of peripheral hearing loss and those that change because of it. To that end, this dissertation provides a framework for beginning to tease out how the relative contributions of peripheral sensory hearing loss (loss of cochlear hair cells) on cortical processing of frequency modulated (FM) sweeps in a mouse model of presbycusis (strain C57/bl6). FM sweeps are sounds that change in frequency over time. Human language and animal vocalizations contain FM sweeps. The genetic engineering tools present in the mouse make it a suitable model for probing the genetic roots and underlying mechanisms of neural dysfunction, such as is seen in presbycusis.

This C57/bl6 mouse strain undergoes predictable high frequency hearing loss by 6 months of age, continuing to profound hearing loss by 12 months of age. The purpose of this dissertation was to establish cortical response properties to FM sweeps as a function of age in a mouse model of presbycusis. Three experiments were performed. In the first experiment, FM sweep response properties were profiled in the young adult core auditory cortex. The main findings of the first experiment indicate that the young adult mouse auditory cortex is selective for a narrow range of FM sweep rates (between 0.5 and 3 kHz/msec) and this selectivity is depth dependent, suggesting FM sweep processing undergoes changes within the a cortical column. The second experiment explored the mechanisms that shape FM sweep rate selectivity. The main findings of the second experiment were that two mechanisms shape FM sweep rate selectivity in the young adult cortex, duration tuning for tones and sideband inhibition. Sideband inhibition is the dominant mechanism in the auditory cortex, shaping FM sweep rate selectivity in approximately 84% of neurons tested. Duration tuning shaped FM sweep rate selectivity in approximately 34% of neurons tested. The third experiment compared direction selectivity, FM rate selectivity, variability, and mechanisms between three age groups, young (1 - 3 months), middle (6-8 months), and old (14 - 20 months). The main findings of the third experiment were that presbycusis results in decreased selectivity, slowed FM rate selectivity, increased variability and decreased reliance on sideband inhibition. The results of these studies serve as a framework for comparisons with mouse strains who do not experience age-related hearing loss.

Fragile X Syndrome (FXS) is a genetic neurodevelopmental disorder that causes autism and intellectual disabilities: exhibiting hyperactivity, elevated anxiety, and impaired cognitive/sensory processing. These deficits result from mutations in the X-linked gene Fragile X messenger ribonucleoprotein 1 (Fmr1). Fmr1-knock- out (KO) mouse models have shown consistency with observations in humans, displaying seizures and sensory processing deficits. Utilizing Fmr1-KO mice to identify a potential treatment for these symptoms, we administered a drug called 2-chloro-4-((2,5-dimethyl-1-(4-trifluoromethoxy)phenyl)-1H-imidazole-4-yl) ethynyl)pyridine (CTEP) to KO mice and measured behavioral changes. CTEP in- hibits metabotropic glutamate receptor pathways, which are upregulated in FXS. Two types of experiments were run: open field test (OFT) and elevated plus maze (EPM), commonly used to study anxiety and hyperactivity. Experimental mice with higher anxiety depict decreased exploration and more time spent near the arena’s wall or closed arms. We found that CTEP reduces the distance traveled in the OFT across both wild-type (WT) and KO groups, suggesting reduced locomotion. There were no statistically significant differences in time spent in EPM closed arms between WT and KO mice, indicating no treatment of anxiety. These results suggest that more effective intervention is needed to target anxiety deficits related to FXS.

Autism spectrum disorder (ASD) is a developmental disorder characterized by symptoms: intellectual difficulties, motor, and language impairment, altered sensory processing, social impediments, and repetitive behaviors. Only 10-20% ASD have clear genetic causes, and fragile X syndrome (FXS) is one of them. FXS is caused by a lack of fragile X messenger ribonucleoprotein (FMRP) whose expression is transcriptionally silenced due to Frm1 gene hypermethylation. Altered sensory processing across all domains has been widely described in humans with FXS. Most noticeable alterations in auditory processing are auditory hyperactivity and impaired temporal processing. The Fmr1 knockout (KO) mouse is a well characterized animal model for FXS studies which also shows signs of auditory hypersensitivity and impaired temporal processing: increased evoked and induced auditory responses, susceptible to audiogenic seizures (AGS), and reduced phase-locking to temporally modulated sound. The consistent manifestations of auditory hyperactivity and impaired temporal processing between human FXS individuals and Frm1 KO make it possible to evaluate potential FXS treatment with Frm1 KO mice. Previous studies by others implied enhancing serotonin signaling ameliorated auditory hyperactivity, yet they failed to pinpoint the specific serotonin receptor subtypes involved. With a more specific and highly selective post-synaptic serotonin-1A (5-HT1A) receptor agonist, NLX-101, we found the AGS susceptibility in developing Frm1 KO mice is significantly reduced, suggesting enhancing 5-HT1A signaling is effective in reducing auditory hyperactivity at the behavioral level in Frm1 KO mice. Electroencephalography (EEG) data acquired from Frm1 KO mice following NLX-101 treatment showed that enhancing 5-HT1A signaling not only reduced auditory hyperactivity at the network level but also improved temporal processing by increasing consistency of the auditory responses to temporally modulated stimuli. RNAscope results showed mRNA transcripts of 5-HT1A receptors are predominately associated with non-GAD cells, implying NLX-101 may reduce auditory hyperactivity in Frm1 KO mice by hyperpolarizing excitatory neurons through 5-HT1A receptor activation.

Autism spectrum disorder (ASD) is currently diagnosed in approximately 1 in 44 children in the United States, based on a wide array of symptoms, including sensory dysfunction and abnormal language development. Auditory temporal processing is crucial for speech recognition and language development. Abnormal development of temporal processing may account for ASD language impairments. Sex differences in the development of temporal processing may underlie the differences in language outcomes in male and female children with ASD. To understand mechanisms of potential sex differences in temporal processing requires a preclinical model. However, there are no studies that have addressed sex differences in temporal processing across development in any animal model of ASD. My dissertation research utilizes two different mouse models of ASD that display sensory abnormalities: the Fmr1 (Fragile X Messenger Ribonucleoprotein 1) knock-out (KO) mouse model of fragile X syndrome (FXS) and a Pten-deletion (phosphatase and tensin homolog missing on chromosome 10) model of autism. I utilized a 40 Hz gap-in-noise ASSR (auditory steady state response) paradigm and in vivo electrophysiology to measure the cortex's reliability in phase locking to brief gaps in noise at varying modulation depths to assess temporal processing acuity. The Fmr1 study results show gap-ASSR deficits in the frontal (FC), but not auditory (AC), cortex in early development (p21, “postnatal day 21”) in male and female KO mice. Unlike male KO mice, female KO mice show wild type-like temporal processing at p30. This research also utilizes a conditional PTEN KO model that displays a specific gene KO pattern, occurring exclusively in the hippocampus and layers III-V of the cortex. Results show that abnormal PTEN expression significantly impacts oscillatory activity in male and female mice, including abnormal resting power distributions and decreased phase-locking compared to controls. Overall, deficits become more prominent in adult mice. Notably, in both ASD models, female KO mice showed increased auditory hypersensitivity compared to males, reflected as increased ERP amplitudes. This dissertation research has identified similarities and differences between two different autism models. Further unraveling these circuits may be the key to understanding and treating ASD pathology.

Fragile X syndrome (FXS) is the most common cause of inherited intellectual impairment and best understood single cause autism. Symptoms of FXS include social anxiety, repetitive behavior, communication disorders, hyperactivity, and seizures. Additionally, event related potential studies using sound stimuli indicate that auditory processing is abnormal in FXS. Many symptoms of FXS have been replicated in the Fmr1 knockout (KO) mice, which serve as a useful model for studying sensory processing abnormalities and symptoms associated with FXS. Fmr1 KO mice exhibit acoustic hypersensitivity and propensity for audiogenic seizures, which suggests altered auditory processing, though the nature of such changes are unknown. Although evidence of unusual social interaction and repetitive behaviors has been found in Fmr1 KO mice, communication anomalies have not yet been modeled. In order to establish the Fmr1 KO mouse as a model for the auditory processing and communication disorders associated with FXS, we developed an assay for assessing vocalization production and performed single unit extracellular electrophysiological recordings in the auditory cortex of Fmr1 KO mice. To quantify possible communication abnormalities in Fmr1 KO mice, we elicited mouse ultrasonic vocalizations (USV) by placing male mice in contact with female mice. This technique revealed that Fmr1 KO mice produce USVs at a decreased rate when compared to wild type mice. Minocycline is a tetracycline analog that has been shown to rescue some symptoms of FXS in open label human trials. Using our USV assay, we demonstrate that minocycline treatment, USV production rate was restored to wild types levels in Fmr1 KO mice. To investigate auditory processing anomalies in Fmr1 KO mice, we performed extracellular single unit recordings in the auditory cortex of anesthetized mice. Presenting single frequency tones revealed expanded frequency tuning, enhanced response magnitude, and greater variability in first spike latency. Frequency modulated sweep stimuli revealed altered sensitivity to FM sweep rate.

Neurodevelopmental disorders (NDD) are a large body of disorders that together affect 1 in 6 children in the U.S. (Center for Disease Control and Prevention). They are characterized by deficits in communication, cognition, attention, socialization and behavior. Abnormalities in sensory processing are also prevalent, and may underlie higher-order processing such as multisensory integration and perception. Determining a root cause in any one NDD is conflated by the multiplicity of brain regions affected, divergence of behavioral outcome measures in animal models and humans, and the multiple mechanistic underpinnings.

Targeting basic sensory processing in NDD is an approach that may address some of these problems. Sensory processing deficits, including auditory, somatosensory, motor and visual, are common in NDDs. While sensory processing may seem far removed from a concept like behavior or socialization, basic sensory processing does serve as the foundation for cognition. Sensory information is relayed to multisensory and emotion-regulation regions of the brain which allow organisms to associate emotional states and salience to sensory information. Learning and memory events such as operant or appetitive conditioning induce stable modifications of processing in sensory regions of the brain, while loss of sensory function can affect working memory and attention. This indicates that disordered sensory processing in fact can affect an organism on multiple levels of function. Understanding the development of basic sensory processing and targeting specific developmental mechanisms may rescue processing deficits in NDD, with the potential for improvements in perception and ‘higher-order’ cognition.

In this series of studies, I focused on the auditory system and sought to increase our understanding of auditory cortex development, and its implications for adult behaviors in a fragile X Syndrome (FXS) mouse model. I discovered the developmental expression patterns of an array of proteins in both early and adult auditory cortex. I discovered differential regulation of proteins within sub-regions of auditory cortex in response to developmental plasticity. I further discovered that dysregulation of these same proteins in adult FXS mice impairs memory formation. Results point to a particular role for perineuronal nets in auditory cortex development and in adult plasticity.

Most older adults experience age-related changes to their hearing, either in the form of hearing loss, or in a more subtle form of hearing impairment, causing difficulties understanding speech in acoustically challenging environments. As the population grows older, better solutions will be needed to alleviate age-related hearing deficits. To test new potential therapies for age-related changes to hearing, preclinical models must be identified and characterized. The studies included in this dissertation measure translationally relevant electrophysiological signals from mice at multiple ages, with and without severe hearing loss, to identify age-related changes to cortical activity, both at baseline and in response to acoustic stimuli. Acoustic stimuli were used to test various aspects of auditory processing, including amplitude modulated signal processing, which supports speech comprehension. We found that age-related changes in the responses to the acoustic stimuli selected for this study mirror many of the changes observed in older humans. We observed age-related deficits in responses to more challenging amplitude modulated signals, even in the absence of hearing loss. In mice with hearing loss, we observed evidence of central gain and disrupted temporal processing. Both results, which mirror age-related changes to human hearing, suggest that aging mice are a useful model for auditory processing in older humans. We then evaluated the therapeutic potential of nicotine for treating age-related changes to auditory temporal processing, with promising results. Nicotine appeared to improve temporal processing in old mice, partially reversing age-related, frequency-specific changes in event-related spectral perturbations and amplitude modulation following responses. This dissertation characterizes translationally relevant preclinical measures of age-related changes to auditory processing, with the aim of testing interventions and therapeutics, such as nicotine, for the treatment of age-related auditory temporal processing deficits.