Biology subjects

Murari, K.

Publications and source records attributed to Murari, K..

10 recordsLinked to original sources

bioRxiv2026

Multi omics reveals mesodermal fate bias and enables predictive cell state control in human pluripotent stem cell biomanufacturing

Human induced pluripotent stem cells can retain conventional markers of pluripotency while acquiring environmentally dependent molecular states that may influence their developmental competence. Yet, how biomanufacturing conditions reorganize the interconnected networks governing cell state remains poorly characterized. Here, we first examined compensation of hiPSCs to static aggregation and dynamic agitation by proteomics. We then examined hiPSCs during single-passage culture in stirred-suspension bioprocesses under varying oxygen and agitation conditions. Intracellular metabolomic and transcriptomic profiling revealed distinct responses across culture configuration, hydrodynamic exposure, and oxygenation. Transition into dynamic culture broadly remodeled mitochondrial organization, carbon allocation, mechanotransduction, proteostasis, and developmental regulation. Increasing agitation produced a persistent response involving cellular architecture, growth-factor signaling, genome maintenance, and Epiblast- and lineage-aligned programs, whereas oxygenation elicited a smaller and more transient metabolic and transcriptional response. Through comparison with a human gastrulation reference, we show that these adaptations intersect with natural developmental programs without reproducing coherent epiblast state or lineage commitment. These findings demonstrate that bioprocess conditions can preserve core pluripotent identity while remodeling broader molecular states associated with developmental responsiveness, providing perspective on evaluating and optimizing hiPSC quality in biomanufacturing beyond restricted marker panels.

bioengineering↗

bioRxiv2026

Online characterization of surrogate metrics for metabolic phenotype in human induced pluripotent stem cell bioprocessing

Human induced pluripotent stem cells (hiPSCs) are the most accessible source material for derivation of stem-cell-based therapies at scale. However, a disconnect exists between quality characteristics of phenotype in the pluripotent state, and downstream metrics for efficacy and safety. Bridging this gap is a major challenge. Given hiPSC plasticity, environmental conditioning plays a crucial role in guiding phenotype. This work presents a parallelizable scale-down approach, acquiring real-time data to inform hiPSC phenotype throughout biomanufacturing. We developed an optoelectronic instrumentation suite capable of measuring pH, dissolved oxygen, and cell density as important surrogates for phenotype in a scale-down expansion bioprocess. We were successful in obtaining continuous, integrated parametric data throughout cultivation and estimating metabolic characteristics of hiPSC phenotype. This system functions as a proof-of-concept tool for development of predictive models and monitoring strategies around the elucidation of phenotypic dynamics within hiPSC biomanufacturing. We have demonstrated a feasible open-source multivariate continuous monitoring approach at research scale that combines common process parameters with a scattering measurement against aggregate density. The combination of these parameters enables surrogate measurement of a metric for metabolic phenotype. This contribution emphasizes monitoring how the bioprocess influences variables important in the context of cell state, in broader pursuit of better understanding the link to downstream functionality and global optima in hiPSC biomanufacturing for regenerative medicine.

bioengineering↗

bioRxiv2025

Early developmental characterization of gap-induced prepulse inhibition and habituation in the Fragile X Syndrome mouse model

IntroductionAtypical sensory processing, particularly auditory hypersensitivity, is a common and debilitating phenotype of Fragile X Syndrome (FXS). Electrophysiology studies in FMR1-knockout (KO) mice previously observed hyperexcitability at the auditory cortex (AC), with enhanced neuronal firing to auditory stimuli. Prepulse inhibition (PPI), a behavioral measure of sensorimotor gating, is robustly impaired in FXS individuals. Interestingly, a related paradigm called gap-induced inhibition of the acoustic startle (GPIAS) is mediated by the AC, and a previous study observed decreased GPIAS in mature FMR1-KO mice. Further, habituation is also an important sensory filtering mechanism, which has been reported to be impaired in mature FMR1-KO mice. However, not much is known about GPIAS and habituation in FMR1- KO mice during early development. MethodsWe evaluated GPIAS in male and female FMR1-KO mice at post-natal days 15 (P15), 20 (P20) and 30 (P30). The paradigm consisted of a prepulse stimulus (a gap in a continuous background noise) followed by a startle stimulus, with an inter-stimulus interval of 50 or 100 ms. Habituation was assessed prior to the GPIAS trials, with a series of startle only stimulus. ResultsWe observed a trend for genotype difference in acoustic startle response (ASR) magnitude, particularly in the female mice at P30. No significant genotype differences were noted in GPIAS or latency of ASR. Response duration was significantly increased in the male FMR1-KO mice compared to their WT counterparts during early development. Significant genotype differences were also observed in habituation and sensitization. In terms of development, we observed a significant increase in GPIAS with maturation. Furthermore, we also observed significant changes in the magnitude, response latency and duration of ASR with maturation. Finally, significant sex differences were observed in ASR magnitude and duration. ConclusionOur findings suggest that behavioral responses to auditory stimuli are dynamic during development and differ between females and males, an important consideration for future study design. Additionally, the FMR1-KO mice display habituation deficits during early development, which could be an ideal window for addressing auditory hypersensitivity in FXS.

neuroscience↗

bioRxiv2025

Alterations in Electroencephalography Signals in Female Fragile X Syndrome Mouse Model on a C57Bl/6J Background

BackgroundFragile X Syndrome (FXS), the most common monogenic cause of autism spectrum disorder, arises from FMR1 gene silencing and exhibits pronounced sex differences in prevalence and phenotypic severity. Electroencephalography (EEG) has emerged as a promising translational biomarker for FXS pathophysiology, yet prior research has predominantly focused on male cohorts. In the widely used C57Bl/6J (B6) mouse strain, male Fmr1 knockout (KO) models show increased absolute gamma power at both juvenile and adult stages, which may reflect cortical hyperexcitability. In contrast, little is known about female Fmr1 KO mice, except that they exhibit no gamma alterations in adulthood. This gap hinders understanding of sex-specific neurodevelopmental trajectories of EEG profile in FXS. Leveraging the genetic stability and translational relevance of the B6 strain, this study compares EEG profiles between juvenile female Fmr1 KO and wild-type (WT) B6 mice to address this critical gap. MethodsFrontal-parietal differential EEG was recorded in freely behaving mice using the Open-Source Electrophysiology Recording system for Rodents. Neural activity was analyzed across three recording conditions: in the home cage, light-dark arena, and open field arena. Computed metrics included absolute/relative power, peak alpha frequency, theta-beta ratio, phase-amplitude coupling, amplitude-amplitude coupling, and multiscale entropy to assess signal complexity. ResultsIn all recording conditions, Fmr1 KO mice exhibited reduced absolute power in theta, alpha, and beta frequency bands compared to WT controls. Relative power analysis revealed decreased alpha activity alongside increased gamma-band power, including both low and high gamma, in the KO mice. Cross-frequency coupling was disrupted, with diminished alpha-gamma phase-amplitude coupling. Amplitude-amplitude coupling between theta or alpha and gamma power displayed distinct changes in different recording conditions. Peak alpha frequency and theta-beta ratio were both reduced or unchanged in the KO mice, depending on the recording condition. Finally, EEG signal complexity remained comparable between the two genotypes across the conditions. Behaviorally, KO mice displayed hyper-exploration in the open field test, characterized by increased center time and entries. However, no overall robust correlations between EEG power in different frequency bands and behavioral parameters in the open field test were observed. Discussion and ConclusionOur results demonstrate that juvenile female Fmr1 KO mice on a B6 background exhibit EEG alterations highly consistent with those reported in FXS patients, particularly increased gamma and reduced alpha power. The robust increase in gamma activity reinforces its status as a reliable biomarker across preclinical and clinical studies, while alpha reductions and slowed peak alpha frequency implicate thalamocortical network involvement. Together, these findings highlight the translational value of this model for studying core circuit dysfunctions in FXS.

neuroscience↗

bioRxiv2025

Alterations in auditory midbrain processing is observed in both female andmale mouse model of Fragile X Syndrome

IntroductionSensory processing deficits presenting as auditory hypersensitivity is a common phenotype associated with Fragile X Syndrome (FXS), a leading monogenic cause of intellectual disability and autism spectrum disorder. Auditory hypersensitivity can also be observed in the FMR1-knockout (KO) mice, a well-established mouse model of autism and FXS. FXS is an X-linked disorder that is more prevalent in males compared to females, as a result most auditory and electrophysiology studies are performed in males. Previous in-vivo electrophysiology studies at the inferior colliculus (IC), an essential component of the auditory pathway, in male FMR1-KO mice at post-natal days 14, 21 and 34 (P14, P21 and P34) demonstrated increased neuronal firing, suggesting that the IC could play a role in auditory hypersensitivity. However, very little is known about the role of the central nucleus of the IC (ICc) and auditory hypersensitivity in females. MethodsHere we investigated auditory processing at P20 and P30, representative of early and late developmental stage of auditory development, respectively, at the central nucleus of the IC (ICc) of both female and male FMR1-KO mice, compared with wildtype (WT) animals. ResultsIn-vivo electrophysiology recordings from the ICc neurons of the FMR1-KO mice at both developmental ages demonstrated increased response magnitude measured by spike number to pure tones of varying frequency and amplitude, compared with age- and sex-matched WT animals. In addition, within the FMR1-KO group we also observed significant developmental and sex difference wherein higher response magnitudes were displayed at P20 and in the female mice. Minimum threshold of ICc neuron in the KO mice was significantly decreased at both P20 and P30. The ICc neurons in the KO mice also displayed increased response duration compared to WT animals at both P20 and P30, but significant sex difference was only observed at P30. Our data also indicated that the ICc neurons of both groups displayed weak negative relationship between latency and response magnitude at P20, and at P30 the WT mice showed a stronger relationship only in the female group. In terms of developmental changes, we observed decreasing neuronal firing only in the KO mice between 20- and 30-day old female and male mice. Reduced response duration was observed in 30-day old mice of both sexes of both genotypes. Regarding minimum threshold, we observed a decline between early and late auditory development only in the male mice. Finally, our results also indicated that in the WT mice the reverse relationship between latency and response magnitude became more pronounced and consistent with age, a developmental trend that was absent in both the female and male FMR1-KO mice. DiscussionOverall, our findings demonstrate that auditory processing deficits can also be observed in the female FMR1-KO mice using in-vivo electrophysiology studies, highlighting the importance of including female subjects in future studies. These results also indicate that auditory hypersensitivity can be observed robustly in younger mice, suggesting that the early development stage could be an ideal target for interventions.

neuroscience↗

bioRxiv2024

Changes in Electroencephalography signals in a juvenile female Fragile X Syndrome mouse model

BackgroundFragile X syndrome (FXS) is the leading monogenic cause of Autism. No broadly effective support option currently exists for FXS, and drug development has suffered many failures in clinical trials based on promising preclinical findings. Thus, effective translational biomarkers of treatment outcomes are needed. Recently, electroencephalography (EEG) has been proposed as a translational biomarker in FXS. Recent years have seen an exciting emergence of novel EEG signal analyses from FXS patients. However, there is a notable gap in corresponding analyses conducted on animal models of the disorder. Being X-linked, FXS is more prevalent in males than females, and there exist significant phenotype differences between males and females with FXS. Recent studies involving male FXS participants and rodent models have identified an increase in absolute gamma EEG power, while alpha power is found to be either decreased or unchanged. However, there is not enough research on female FXS patients or models. In addition, studying EEG activity in both young and adult FXS patients or rodent models is crucial for better understanding of the disorders effects on brain development. Therefore, using the well established fmr1 knockout (KO) mouse model of FXS, we aim to compare EEG signal between female wild-type (WT) and female model mice at both juvenile and adult ages. MethodsFrontal-parietal differential EEG was recorded using a stand-alone Open-Source Electrophysiology Recording system for Rodents (OSERR). EEG activity was recorded in three different conditions: a) in the subjects home cage, and in the arenas for b) light -dark test and c) open field test. Absolute and relative EEG power as well as peak alpha frequency, theta-beta ratio, phase-amplitude and amplitude-amplitude coupling, and EEG signal complexity were computed for each condition. ResultsIn our study, we found absolute alpha, beta, gamma and total EEG power is increased in the female model compared to WT controls at the juvenile and adult ages. Alongside, relative theta power is decreased in the model. Additionally, phase-amplitude and amplitude-amplitude coupling is altered in the model. Furthermore, peak alpha frequency is increase, and theta-beta ratio is decreased in the model. Lastly, no change in EEG signal complexity is found. Discussion and ConclusionConsistent with most findings from FXS patients and rodent models, our results demonstrated an increase in gamma power in fmr1KO female mice, reinforcing gamma power as a robust and reliable EEG phenotype across FXS models. Additionally, theta-gamma cross frequency amplitude coupling is inversely coupled in female FXS model, which is similar to what has been reported in FXS patients. Overall, our findings reveal that not all EEG biomarkers observed in FXS patients are replicated in the female FXS model. For example, peak alpha frequency, theta-beta ratio, and brain signal complexity showed discrepancies between the mouse models and FXS patients. Additionally, when compared to previously reported EEG changes in male FXS mouse models, our results highlight the presence of a potential sex-based difference in EEG phenotypes at both juvenile and adult stages of fmr1 KO mouse models. Together, our study indicates that certain EEG parameters may be more translatable between rodent models and FXS patients than others and underscore the importance of considering sex and developmental stage as a critical factor when using EEG as a biomarker in FXS research.

neuroscience↗

bioRxiv2024

Exploring the Dynamics of Social Interactions During the Juvenile Stage in a Mouse Model of Fragile X Syndrome

IntroductionFragile X Syndrome (FXS), caused by Fmr1 mutations, is linked to cognitive and behavioral differences, including altered social interactions. Most mouse studies focus on adults, despite human research showing critical developmental changes in childhood and adolescence. We examined social behavior in juvenile male and female Fmr1 knockout (KO) mice as well as heterozygous (HET) females. We further assessed cortical activity in KO females to better understand early phenotypes. MethodsJuvenile mice of both sexes and genotypes were paired in same-sex, novel dyads for 10-minute interactions. Key social behaviors such as head, anogenital, and body sniffing, and physical touch, as well as distance travelled, were analyzed with a marker-less tracking software. Frontal-parietal EEG recordings were collected from wild-type (WT) and KO females in home cage and social contexts to analyze power spectra across frequency bands. ResultsHET and KO females engaged in more frequent but shorter interaction events compared to WT females, with HET females showing the highest counts. Males displayed similar trends when comparing KO and WT. Males engaged in overall higher interaction events than females. EEG analyses revealed altered oscillatory activity in KO females compared to WT females, especially within theta, alpha, and beta bands, most prominently during the early interaction phase. Locomotor activity correlated weakly with head/anogenital sniffing but more strongly with body sniffing and touch. DiscussionThese findings suggest that Fmr1-related differences in juvenile social behavior are sex-dependent and associated with cortical oscillatory changes. Characterizing these early phenotypes in both sexes allows us to further understand FXS development and informs potential routes for early intervention. Key PointsO_LIFragile X Syndrome (FXS), the leading inherited cause of autism, is associated with disruptions in social behavior. C_LIO_LIWhile social phenotypes are relatively well described in adult mouse models of FXS, juvenile manifestations remain poorly understood. C_LIO_LISocial behavior was assessed in juvenile male and female Fmr1 knockout (KO), heterozygous (HET, female only), and wildtype (WT) mice, and frontal-parietal EEG recordings were collected from WT and KO females. C_LIO_LIHET and KO females exhibited more frequent but shorter social interactions than WT females, with HET showing the greatest number of events. Males showed similar patterns when comparing KO and WT. Males engaged in higher overall interaction events than females. EEG recordings revealed altered oscillatory activity in KO compared to WT females, most pronounced during the early phase of social encounter. C_LIO_LIThese findings reveal sex- and genotype-dependent differences in juvenile social behavior and cortical activity, highlighting the importance of studying juvenile development in FXS. C_LI

animal behavior and cognition↗

bioRxiv2023

Improving vocal communication with a ketogenic diet in a mouse model of autism

BackgroundDeficits in social communication and language development is a hallmark of autism spectrum disorder currently with no cure. Interventional studies using animal models have been very limited in demonstrating improved vocal communication. Autism has been proposed to involve metabolic dysregulation. Ketogenic diet (KD) is a metabolism-based therapy for medically intractable epilepsy, and its applications in other neurological conditions have been increasingly tested. However, how it would affect vocal communication has not been explored. The BTBR mouse strain is considered a model of idiopathic autism. They display robust deficits in vocalization during social interaction, and have metabolic changes implicated in autism. MethodsWe investigated the effects of KD on ultrasonic vocalizations (USVs) in juvenile and adult BTBR mice during male-female social encounters. ResultsAfter a brief treatment with KD, the amount, spectral bandwidth, and much of the temporal structure of USVs were robustly improved in both juvenile and adult BTBR mice. Composition of call categories and transitioning between individual call subtypes was more effectively improved in juvenile BTBR mice. LimitationsAlthough sharing certain attributes, mouse vocalization is unlikely to model all aspects in the development and deficits of human language. KD is highly restrictive and can be difficult to administer, especially for many people with autism who have narrow food selections. Side effects and potential influence on development should also be considered. Future studies are required to tease apart the molecular mechanisms of KDs effects on vocalization. ConclusionsTogether, our data provide further support to the hypothesis that metabolism-based dietary intervention could modify disease expression, including core symptoms, in autism.

animal behavior and cognition↗

bioRxiv2021

Markerless mouse tracking for social experiments

Automated behavior quantification requires accurate tracking of animals. Simultaneous tracking of multiple animals, particularly those lacking visual identifiers, is particularly challenging. Problems of mistaken identities and lost information on key anatomical features are common in existing methods. Here we propose a markerless video-based tool to simultaneously track two socially interacting mice of the same appearance. It incorporates conventional handcrafted tracking and deep learning based techniques, which are trained on a small number of labeled images from a very basic, uncluttered experimental setup. The output consists of body masks and coordinates of the snout and tail-base for each mouse. The method was tested on a series of cross-setup videos recorded under commonly used experimental conditions including bedding in the cage and fiberoptic or headstage implants on the mice. Results obtained without any human intervention showed the effectiveness of the proposed approach, evidenced by a near elimination of identities switches and a 10% improvement in tracking accuracy over a pure deep-learning-based keypoint tracking approach trained on the same data. Finally, we demonstrated an application of this approach in studies of social behaviour of mice, by using it to quantify and compare interactions between pairs of mice in which some are anosmic, i.e. unable to smell. Our results indicated loss of olfaction impaired typical snout-directed social recognition behaviors of mice, while non-snout-directed social behaviours were enhanced. Together, these results suggest that the hybrid approach could be valuable for studying group behaviors in rodents, such as social interactions.

bioengineering↗

bioRxiv2021

Spatiotemporal components of sustained functional hyperemia are differentially modulated by locomotion and silenced with vascular chemogenetics

Neural activity underlying sensation, movement or cognition drives regional blood flow enhancement - termed functional hyperemia - to increase the oxygen supply to respiring cells for as long as needed to meet energy demands. However, functional hyperemia is often studied under anesthesia which typically yields response profiles that appear temporally and spatially homogenous. We have insufficient understanding of the underlying kinetics of oxygen delivery in awake animals, especially during specific behaviours that may influence neurally-driven enhancements in cerebral blood flow. Using widefield intrinsic optical signal imaging in awake, head-fixed but active mice, we demonstrated distinct early and late components to changes in intravascular oxygenation in response to sustained (30s) whisker stimulation. We found that the late component (20-30s), but not the early component (1-5s), was strongly influenced by level of whisking/locomotion in the region of highest response and in surrounding regions. Optical flow analyses revealed complex yet stereotyped spatial properties of the early and late components that were related to location within the optical window and the initial state of the cerebral vasculature. In attempt to control these complex response characteristics, we drove a canonical microvasculature constriction pathway using mural cell Gq-chemogenetic mice. A low-dose of systemic C21 strongly limited both the magnitude and spatial extent of the sensory-evoked hemodynamic response, showing that functional hyperemia can be severely limited by direct mural cell activation. These data provide new insights into the cerebral microcirculation in the awake state and may have implications for interpreting functional imaging data.

neuroscience↗