Biology subjects

Odierna, L.

Publications and source records attributed to Odierna, L..

2 recordsLinked to original sources

bioRxiv2024

Stable primary brain cell cultures from zebrafish reveal hyperproliferation of non-neuronal cells from scn1lab mutants

Zebrafish are a popular model system for studying the genetic and neural underpinnings of perception and behavior, both in wild-type animals and in the context of disease modelling. Cultured primary neurons provide a key complementary tool for such studies, but existing protocols for culturing embryonic zebrafish primary neurons are limited by short cell survival and low neuronal purity. In this study, we set out to establish a protocol to produce long lived, pure neuronal cultures from zebrafish that could be used to study the mechanistic contributions of genes to neuronal networks. We then used these primary cultures to characterize cell proliferation and differentiation in primary neurons derived from scn1lab mutant embryos, which lack a sodium channel relevant to Dravet syndrome and autism. Using our optimized protocol, we generated cultures that proliferate, diversify, and form stable networks of neurons surviving for months. These stable cultures allowed us to perform genetic experiments, in this case revealing dramatic differences in the cellular composition of cultures derived from scn1lab mutant embryos versus their wild type siblings. Specifically, we find that loss of scn1lab promotes hyperproliferation of non-neuronal cells in mixed cultures of brain cells. In pure neuronal cultures, we find alterations in neurotransmitter subtypes consistent with known effects of scn1lab loss of function. Validating the utility of this approach, we then identify a corresponding hyperproliferation phenotype in live scn1lab mutant embryos, shedding light on potential mechanisms that may be relevant for Dravet syndrome. Significance statementMost existing embryonic zebrafish primary neuron culture protocols describe growing mixed cell types for restricted durations. Here, we report generation of zebrafish mixed type or pure neuronal cultures that are viable for over 100 days. We apply these cultures to gain new insight into scn1lab, a zebrafish orthologue of the Dravet Syndrome-associated sodium channel gene SCN1A. We report that loss of scn1lab results in hyperproliferation of non-neuronal cells, revealing an underappreciated mechanism by which mutations in SCN1A impact the structure and function of neuronal networks. Our in vitro cultures thus faithfully recapitulate in vivo neurobiology and provide a powerful platform to interrogate brain function in health and disease.

neuroscience↗

bioRxiv2024

Biological sex determines skeletal muscle atrophy in response to cortical TDP-43 pathology

BackgroundAmyotrophic lateral sclerosis (ALS) is a fatal and incurable neurodegenerative condition. In ALS, wasting of skeletal muscle causes weakness, paralysis and ultimately, death due to respiratory failure. Diagnosis of ALS is a long process and delays in diagnosis are common, which impedes rapid provision of patient care and treatment. Additional tools or methodologies that improve early detection might help overcome the diagnostic delays and enhance survival and quality of life for people with ALS. In this study, we used a transgenic mouse model to create a detailed catalogue of skeletal muscle wasting with the goal of finding muscles that can be examined to enhance early diagnosis of ALS. MethodsCortical pathology was induced by crossing CaMKIIa-tTA and tetO-hTDP-43{Delta}NLS transgenic mice ({Delta}NLS). Transgenic expression was induced at 30-days postnatal via removal of doxycycline diet. Mice were aged to 15-, 20-, 30- and 45-days post transgene induction. Microdissection was applied to isolate 22 individual hindlimb muscles for measurement of weight. Both males and females were used at all timepoints. ResultsWe found that male and female {Delta}NLS mice exhibited hindlimb skeletal muscle atrophy relative to controls. Multiply innervated muscles, also known as series-fibered muscles, were especially vulnerable to atrophy. The strongest predictor of the atrophic response across all hindlimb muscles was the extent to which any individual muscle was larger in males than females, known also as sexual dimorphism. In males, muscles that are usually larger in males compared to females experienced the most atrophy. Conversely, in females, muscles that are usually of similar size between males and females experienced the most atrophy. Segregating muscles based on whether they were more affected in males or females revealed that hip extensors, knee flexors, knee extensors, ankle dorsiflexors and ankle evertors were more affected in males. Hip adductors, hip rotators, hip flexors and ankle plantarflexors were more affected in females. ConclusionsOur results demonstrate that the difference in the size of skeletal muscles in males compared to females is the most powerful predictor of muscle atrophy in response to dying forward pathology. This indicates that sex is a strong determinant of skeletal muscle vulnerability in ALS. Our results provide new insights into determinants of skeletal muscle atrophy and may help inform selection of muscles for diagnostic testing of ALS patients.

neuroscience↗