Patient-specific mutation of Dync1h1 in mice causes brain and behavioral deficits.

AIMS: Cytoplasmic dynein heavy chain (DYNC1H1) is a multi-subunit protein complex that provides motor force for movement of cargo on microtubules and traffics them back to the soma. In humans, mutations along the DYNC1H1 gene result in intellectual disabilities, cognitive delays, and neurologic and motor deficits. The aim of the study was to generate a mouse model to a newly identified de novo heterozygous DYNC1H1 mutation, within a functional ATPase domain (c9052C > T(P3018S)), identified in a child with motor deficits, and intellectual disabilities. RESULTS: P3018S heterozygous (HET) knockin mice are viable; homozygotes are lethal. Metabolic and EchoMRI™ testing show that HET mice have a higher metabolic rate, are more active, and have less body fat compared to wildtype mice. Neurobehavioral studies show that HET mice perform worse when traversing elevated balance beams, and on the negative geotaxis test. Immunofluorescent staining shows neuronal migration abnormalities in the dorsal and lateral neocortex with heterotopia in layer I. Neuron-subtype specific transcription factors CUX1 and CTGF identified neurons from layers II/III and VI respectively in cortical layer I, and abnormal pyramidal neurons with MAP2+ dendrites projecting downward from the pial surface. CONCLUSION: The HET mice are a good model for the motor deficits seen in the child, and highlights the importance of cytoplasmic dynein in the maintenance of cortical function and dendritic orientation relative to the pial surface. Our results are discussed in the context of other dynein mutant mice and in relation to clinical presentation in humans with DYNC1H1 mutations.

Knockout of the intellectual disability-linked gene Hs6st2 in mice decreases heparan sulfate 6-O-sulfation, impairs dendritic spines of hippocampal neurons, and affects memory.

Heparan sulfate (HS) is a linear polysaccharide that plays a key role in cellular signaling networks. HS functions are regulated by its 6-O-sulfation, which is catalyzed by three HS 6-O-sulfotransferases (HS6STs). Notably, HS6ST2 is mainly expressed in the brain and HS6ST2 mutations are linked to brain disorders, but the underlying mechanisms remain poorly understood. To determine the role of Hs6st2 in the brain, we carried out a series of molecular and behavioral assessments on Hs6st2 knockout mice. We first carried out strong anion exchange-high performance liquid chromatography and found that knockout of Hs6st2 moderately decreases HS 6-O-sulfation levels in the brain. We then assessed body weights and found that Hs6st2 knockout mice exhibit increased body weight, which is associated with abnormal metabolic pathways. We also performed behavioral tests and found that Hs6st2 knockout mice showed memory deficits, which recapitulate patient clinical symptoms. To determine the molecular mechanisms underlying the memory deficits, we used RNA sequencing to examine transcriptomes in two memory-related brain regions, the hippocampus and cerebral cortex. We found that knockout of Hs6st2 impairs transcriptome in the hippocampus, but only mildly in the cerebral cortex. Furthermore, the transcriptome changes in the hippocampus are enriched in dendrite and synapse pathways. We also found that knockout of Hs6st2 decreases HS levels and impairs dendritic spines in hippocampal CA1 pyramidal neurons. Taken together, our study provides novel molecular and behavioral insights into the role of Hs6st2 in the brain, which facilitates a better understanding of HS6ST2 and HS-linked brain disorders.

Mustn1 ablation in skeletal muscle results in functional alterations.

Mustn1, a gene expressed exclusively in the musculoskeletal system, was shown in previous in vitro studies to be a key regulator of myogenic differentiation and myofusion. Other studies also showed Mustn1 expression associated with skeletal muscle development and hypertrophy. However, its specific role in skeletal muscle function remains unclear. This study sought to investigate the effects of Mustn1 in a conditional knockout (KO) mouse model in Pax7 positive skeletal muscle satellite cells. Specifically, we investigated the potential effects of Mustn1 on myogenic gene expression, grip strength, alterations in gait, ex vivo investigations of isolated skeletal muscle isometric contractions, and potential changes in the composition of muscle fiber types. Results indicate that Mustn1 KO mice did not present any substantial phenotypic changes or significant variations in genes related to myogenic differentiation and fusion. However, an approximately 10% decrease in overall grip strength was observed in the 2-month-old KO mice in comparison to the control wild type (WT), but this decrease was not significant when normalized by weight. KO mice also generated approximately 8% higher vertical force than WT at 4 months in the hindlimb. Ex vivo experiments revealed decreases in about 20 to 50% in skeletal muscle contractions and about 10%-20% fatigue in soleus of both 2- and 4-month-old KO mice, respectively. Lastly, immunofluorescent analyses showed a persistent increase of Type IIb fibers up to 15-fold in the KO mice while Type I fibers decreased about 20% and 30% at both 2 and 4 months, respectively. These findings suggest a potential adaptive or compensatory mechanism following Mustn1 loss, as well as hinting at an association between Mustn1 and muscle fiber typing. Collectively, Mustn1's complex roles in skeletal muscle physiology requires further research, particularly in terms of understanding the potential role of Mustn1 in muscle repair and regeneration, as well as with influence of exercise. Collectively, these will offer valuable insights into Mustn1's key biological functions and regulatory pathways.

Neocortical and cerebellar malformations affect flurothyl-induced seizures in female C57BL/6J mice.

Brain malformations cause cognitive disability and seizures in both human and animal models. Highly laminated structures such as the neocortex and cerebellum are vulnerable to malformation, affecting lamination and neuronal connectivity as well as causing heterotopia. The objective of the present study was to determine if sporadic neocortical and/or cerebellar malformations in C57BL/6J mice are correlated with reduced seizure threshold. The inhaled chemi-convulsant flurothyl was used to induce generalized, tonic-clonic seizures in male and female C57BL/6J mice, and the time to seizure onset was recorded as a functional correlate of brain excitability changes. Following seizures, mice were euthanized, and brains were extracted for histology. Cryosections of the neocortex and cerebellar vermis were stained and examined for the presence of molecular layer heterotopia as previously described in C57BL/6J mice. Over 60% of mice had neocortical and/or cerebellar heterotopia. No sex differences were observed in the prevalence of malformations. Significantly reduced seizure onset time was observed dependent on sex and the type of malformation present. These results raise important questions regarding the presence of malformations in C57BL/6J mice used in the study of brain development, epilepsy, and many other diseases of the nervous system.

Haploinsufficiency of Shank3 in Mice Selectively Impairs Target Odor Recognition in Novel Background Odors.

Individuals with mutations in a single copy of the SHANK3 gene present with social interaction deficits. Although social behavior in mice depends on olfaction, mice with mutations in a single copy of the Shank3 gene do not have olfactory deficits in simple odor identification tasks (Drapeau et al., 2018). Here, we tested olfaction in mice with mutations in a single copy of the Shank3 gene (Peça et al., 2011) using a complex odor task and imaging in awake mice. Average glomerular responses in the olfactory bulb of Shank3B +/- were correlated with WT mice. However, there was increased trial-to-trial variability in the odor responses for Shank3B +/- mice. Simulations demonstrated that this increased variability could affect odor detection in novel environments. To test whether performance was affected by the increased variability, we tested target odor recognition in the presence of novel background odors using a recently developed task (Li et al., 2023). Head-fixed mice were trained to detect target odors in the presence of known background odors. Performance was tested using catch trials where the known background odors were replaced by novel background odors. We compared the performance of eight Shank3B +/- mice (five males, three females) on this task with six WT mice (three males, three females). Performance for known background odors and learning rates were similar between Shank3B +/- and WT mice. However, when tested with novel background odors, the performance of Shank3B +/- mice dropped to almost chance levels. Thus, haploinsufficiency of the Shank3 gene causes a specific deficit in odor detection in novel environments. Our results are discussed in the context of other Shank3 mouse models and have implications for understanding olfactory function in neurodevelopmental disorders.SIGNIFICANCE STATEMENT People and mice with mutations in a single copy in the synaptic gene Shank3 show features seen in autism spectrum disorders, including social interaction deficits. Although mice social behavior uses olfaction, mice with mutations in a single copy of Shank3 have so far not shown olfactory deficits when tested using simple tasks. Here, we used a recently developed task to show that these mice could identify odors in the presence of known background odors as well as wild-type mice. However, their performance fell below that of wild-type mice when challenged with novel background odors. This deficit was also previously reported in the Cntnap2 mouse model of autism, suggesting that odor detection in novel backgrounds is a general deficit across mouse models of autism.

Mustn1 ablation in skeletal muscle results in increased glucose tolerance concomitant with upregulated GLUT expression in male mice.

Glucose homeostasis is closely regulated to maintain energy requirements of vital organs and skeletal muscle plays a crucial role in this process. Mustn1 is expressed during embryonic and postnatal skeletal muscle development and its function has been implicated in myogenic differentiation and myofusion. Whether Mustn1 plays a role in glucose homeostasis in anyway remains largely unknown. As such, we deleted Mustn1 in skeletal muscle using a conditional knockout (KO) mouse approach. KO mice did not reveal any specific gross phenotypic alterations in skeletal muscle. However, intraperitoneal glucose tolerance testing (IPGTT) revealed that 2-month-old male KO mice had significantly lower glycemia than their littermate wild type (WT) controls. These findings coincided with mRNA changes in genes known to be involved in glucose metabolism, tolerance, and insulin sensitivity; 2-month-old male KO mice had significantly higher expression of GLUT1 and GLUT10 transporters, MUP-1 while OSTN expression was lower. These differences in glycemia and gene expression were statistically insignificant after 4 months. Identical experiments in female KO and WT control mice did not indicate any differences at any age. Our results suggest a link between Mustn1 expression and glucose homeostasis during a restricted period of skeletal muscle development/maturation. While this is an observational study, Mustn1's relationship to glucose homeostasis appears to be more complex with a possible connection to other key proteins such as GLUTs, MUP-1, and OSTN. Additionally, our data indicate temporal and sex differences. Lastly, our findings strengthen the notion that Mustn1 plays a role in the metabolic capacity of skeletal muscle.

Robust odor identification in novel olfactory environments in mice.

Relevant odors signaling food, mates, or predators can be masked by unpredictable mixtures of less relevant background odors. Here, we developed a mouse behavioral paradigm to test the role played by the novelty of the background odors. During the task, mice identified target odors in previously learned background odors and were challenged by catch trials with novel background odors, a task similar to visual CAPTCHA. Female wild-type (WT) mice could accurately identify known targets in novel background odors. WT mice performance was higher than linear classifiers and the nearest neighbor classifier trained using olfactory bulb glomerular activation patterns. Performance was more consistent with an odor deconvolution method. We also used our task to investigate the performance of female Cntnap2-/- mice, which show some autism-like behaviors. Cntnap2-/- mice had glomerular activation patterns similar to WT mice and matched WT mice target detection for known background odors. However, Cntnap2-/- mice performance fell almost to chance levels in the presence of novel backgrounds. Our findings suggest that mice use a robust algorithm for detecting odors in novel environments and this computation is impaired in Cntnap2-/- mice.

Neurodevelopmental malformations of the cerebellum and neocortex in the Shank3 and Cntnap2 mouse models of autism.

There are many mouse models of autism with broad use in neuroscience research. Genetic background can be a major contributor to the phenotype observed in any mouse model of disease, including genetic models of autism. C57BL/6 mice display spontaneous glio-neuronal heterotopia in the cerebellar vermis and neocortex which may also exist in mouse models of autism created on this background. In the present report, we document the presence of cerebellar and neocortical heterotopia in heterozygous and KO Shank3 and Cntnap2 mice which are due to the C57BL/6 genotype and discuss the role these malformations may play in research using these genetic models of autism.

Undergraduate Participation in the Society for Neuroscience.

Undergraduate neuroscience education has grown substantially in the US as well as participation in outreach and research activities by undergraduates. In line with these observations, undergraduates may also be seeking membership in the Society for Neuroscience (SfN) as well as attending the SfN annual meeting. Data in the present report show that undergraduate membership in SfN has increased between 2013 and 2019 as well as annual undergraduate SfN meeting attendance and abstract submissions for research presentations. Increases were observed for both US and international undergraduates. These data are discussed in the context of motivations of undergraduates to pursue future academic training and/or careers in neuroscience. These data are important to faculty and administrators at institutions that currently have or seek to establish new undergraduate neuroscience programs given membership in a professional society and attendance at a major conference can positively impact academic and professional development.

Primary Literature In Clinical Neuroscience for In-Person Or Remote Instruction.

Structure and function relationships in the nervous system are a major component of neuroscience education. Readings and/or discussion of lesion studies in animal models are often used to demonstrate how brain injury/damage affects specific behaviors or cognitive processes. In contrast, primary literature in clinical neuroscience is less often used to teach brain structure and function relationships and this literature often describes remarkable stories of preserved brain function despite major brain injury/lesion. Here we describe a series of published articles in clinical neuroscience that we used in an undergraduate neuroscience course that challenge the simplistic views of brain localization of function and demonstrate the dynamic and plastic properties of the brain. Discussion of these primary articles can take place in-person or remote via video conferencing platforms.

Combination of acute intravenous methamphetamine injection and LPS challenge facilitate leukocyte infiltration into the central nervous system of C57BL/6 mice.

Methamphetamine (METH) is a stimulant of the central nervous system (CNS) that causes behavioral changes in users. METH is slowly cleared from brain tissue and its chronic use is neurotoxic. METH also alters the cellular and chemical components of inflammation. However, little is known about the effect of a single intravenous dose of METH followed by bacterial lipopolysaccharide (LPS) injection on cellular infiltration and cytokine release in brain tissue. Using a murine model of acute METH administration and flow cytometry, we found that combination of METH and LPS stimulate the infiltration of macrophages (F4/80+cells) and neutrophils (Ly-6G+cells) into the CNS. Histological sections of the brainstem of METH-treated and LPS-challenged C57BL/6 mice demonstrated considerable leukocyte infiltration relative to untreated, LPS, and METH groups. Moreover, rodents treated with LPS alone or combined with METH showed elevated levels of pro-inflammatory cytokines mRNA in brain tissue. Our observations are important because recognizing neuroinflammatory changes after acute METH administration might help us to understand METH-induced neurotoxicity in users.

An Ultrasonically Powered Wireless System for In Vivo Gastric Slow-Wave Recording.

This paper summarizes our recent progress towards Gastric Seed which is an ultrasonically interrogated millimeter-sized implant for gastric electrical activity (also known as slow waves, SWs) recording. We present a proof-of-concept wireless system designed to collect, transmit, and store in vivo SW signals by integrating a prototype Gastric Seed chip, fabricated in a 0.35-µm 2P4M CMOS process, with a commercial-off-the-shelf (COTS) amplifier, 10-bit analog-to-digital converter (ADC), and pair of microcontrollers (MCU) as radio-frequency (RF) transceivers. The chip includes ultrasonic self-regulated power management and addressable pulse-based data transfer. Utilizing two pairs of millimeter-sized stacked power/data ultrasonic transducers spaced by 6 cm in a water tank, the chip achieved a regulated voltage of 2.5 V and a data rate of 16 kbps. The amplifier was configured to have a gain of 60 dB with a 3-dB bandwidth of 18 mHz to 500 mHz. The MCU's built-in 10-bit ADC and RF transceiver were used to digitize the SW signal and transmit the data to a computer. In vivo, SW was recorded wirelessly from the stomach of an anesthetized rat. The recorded SWs showed a frequency of 1.5 cycle-per-minute (cpm) and maximum and minimum amplitudes of 1.03 mV and 0.28 mV peak-to-peak, respectively.

Quantitative Indicators of Continued Growth in Undergraduate Neuroscience Education in the US.

There is both anecdotal and quantitative evidence that undergraduate neuroscience education has grown substantially in the US. Therefore, efforts to continue to track changes in undergraduate neuroscience education are important. Here we provide quantitative data that both public and private institutions are creating new undergraduate neuroscience programs. In addition, we demonstrate that the number of graduates from undergraduate neuroscience programs continues to increase compared to graduates from other life sciences programs. These data are important to faculty and administrators at institutions that currently have or seek to establish new undergraduate neuroscience programs.