Biology subjects

O'Bryan, M.

Publications and source records attributed to O'Bryan, M..

7 recordsLinked to original sources

bioRxiv2025

Acute Stress in Female Adolescent Rats Increases Anxiety-like but not Depression-like Behaviors

Adolescence is a critical developmental period where heightened stress reactivity and responsiveness increase susceptibility to psychiatric conditions like anxiety and depression. Studies with rodent models have primarily tested the effects of chronic, rather than acute, stress on the development of anxiety- and depression-like behaviors, and many have not included female subjects. The current study examined whether a single acute stress exposure in female rats can induce anxiety-like and depression-like behaviors, and whether the timing of the stress, early (postnatal day 28) versus mid-adolescence (postnatal day 36), influences these outcomes. We used a PTSD-relevant stress paradigm that paired acute immobilization with fox-urine predator odor. Female Sprague-Dawley rats completed a battery of behavioral assays to characterize anxiety-like (elevated plus maze, open field test) and depression-like behaviors (sucrose preference test, forced swim test) between 12-16 days post exposure. Acute stress in either early or mid-adolescence caused an increase in anxiety-like behavior on the elevated plus maze 15 days post-exposure. No increase in depression-like behaviors was observed at either age. Our findings suggest that a single exposure to a predator odor when paired with immobilization is sufficient to increase anxiety-like but not depression-like behaviors in female adolescent rats. These results provide evidence that acute stress during either early or mid-adolescence can lead to later anxiety in females.

animal behavior and cognition↗

bioRxiv2025

Position-dependent effects of SCN2A premature stop codons on neuronal excitability and behavior

SCN2A encodes the voltage-gated sodium channel NaV1.2, a key determinant of spike initiation and propagation in glutamatergic neurons. Premature termination codons are often assumed to produce uniform haploinsufficiency via nonsense-mediated decay, yet whether distinct SCN2A premature stop codons yield equivalent molecular, cellular, and behavioral outcomes remains unknown. We generated two mouse lines carrying patient mutations--Scn2aY84X/+ (p.Tyr84UAA; early coding sequence) and Scn2aR1627X/+(p.Arg1627UGA; terminal coding exon)--on a C57BL/6J background. Allele-specific expression was quantified by targeted next-generation sequencing of whole-brain reverse transcribed cDNA. NaV1.2 protein was measured in half-brain lysates by automated western blot and ex vivo whole-cell recordings were obtained from layer 5b pyramidal-tract neurons in medial prefrontal cortex. A panel of behavioral assays assessed locomotion/exploration, motor learning, anxiety-like behavior, sociability, sensorimotor gating, and seizure susceptibility. Allele-specific RNA handling diverged by position: mRNA carrying Y84X engaged partial nonsense-mediated decay, whereas R1627X transcripts were at allelic balance. Despite this difference in RNA fate, NaV1.2 protein was comparably reduced in both lines. Electrophysiologically, both premature termination codon mutations slowed the action-potential upstroke, with a larger decrement in Scn2aY84X/+than in Scn2aR1627X/+. Spike threshold was depolarized only in Scn2aY84X/+, whereas Scn2aR1627X/+ remained similar to wild type. Frequency-current relations showed reduced firing at near-rheobase inputs in both mutants, with responses approaching wild type at stronger currents. Behaviorally, locomotion, sociability, and sensorimotor gating were preserved. Both lines exhibited increased grooming--consistent with restrictive, repetitive behavior; Scn2aY84X/+alone showed greater exploration in the elevated-risk context and a male-predominant deficit in rotarod learning. In maximal electroshock testing, mortality was lower in both lines without differences in seizure threshold or severity. Our results show that distinct SCN2A premature termination codons are not equivalent to one another, nor to a uniform haploinsufficient state. An early, nonsense-mediated decay-competent premature stop codon (Y84X) and a terminal-exon one (R1627X) produce partially overlapping yet allele-specific effects on neuronal excitability and behavior. These findings establish premature termination codon position as a determinant of phenotype, supporting allele-tailored mechanistic studies and therapeutic strategies.

neuroscience↗

bioRxiv2023

Fertility is compromised after oocyte-specific deletion of Katanin A-subunit, Katna1, but not Katnal1

Katanins are microtubule severing enzymes that play roles in shaping diverse microtubule-based structures during all cell cycle stages. To address the role of katanin A-subunits in mammalian oocytes, we have used the Zp3-CreLox approach to specifically delete katanin A1 (Katna1) and katanin A-like 1 (Katnal1) from the start of oocyte growth in mice. Here, we show that Katnal1 is not required for normal female fertility, but that deletion of Katna1 causes a 50% decrease in fertility. Further investigation in Katna1-/- oocytes revealed no effect on MI spindle morphology but a significant effect on the morphology of MII spindles. This was accompanied by a decreased rate of fertilisation. Resultant Katna1+/- heterozygous embryos that reached the 2-cell stage developed at normal rates to the blastocyst stage. Diploid homozygous parthenotes derived from Katna1-/- oocytes revealed a reduced rate of blastocyst formation, decreased cell number and increased nuclear size. The ability of the paternal allele to rescue preimplantation development suggests the origin of the decrease in the fertility of conditional Katna1-/- mice lies in abnormalities arising in the egg to embryo transition prior to embryonic genome activation.

developmental biology↗

bioRxiv2022

Male mammalian meiosis and spermiogenesis is critically dependent on the shared functions of the katanins KATNA1 and KATNAL1

Katanin microtubule severing enzymes are potent M-phase regulators in oocytes and somatic cells. How the complex, and evolutionarily critical, male mammalian meiotic spindle is sculpted remains unknown. Here, using multiple single and double gene knockout mice, we reveal that the canonical katanin A-subunit, KATNA1, and its close paralogue, KATNAL1, together execute multiple aspects of meiosis. We show KATNA1 and KATNAL1 collectively regulate the male meiotic spindle, cytokinesis and midbody abscission, in addition to diverse spermatid remodelling events, including Golgi organisation, and acrosome and manchette formation. We also define KATNAL1-specific roles in sperm flagella development, manchette regulation, and sperm-epithelial disengagement. Finally, using proteomic approaches we define the KATNA1, KATNAL1, and KATNB1 mammalian testis interactome, which includes a network of cytoskeletal and vesicle trafficking proteins. Collectively, we reveal the presence of multiple katanin A-subunit paralogs in mammalian spermatogenesis allows for customized cutting via neofunctionalization and protective buffering via gene redundancy.

cell biology↗

bioRxiv2021

A de novo paradigm for male infertility

IntroductionDe novo mutations (DNMs) are known to play a prominent role in sporadic disorders with reduced fitness1. We hypothesize that DNMs play an important role in male infertility and explain a significant fraction of the genetic causes of this understudied disorder. To test this hypothesis, we performed trio-based exome-sequencing in a unique cohort of 185 infertile males and their unaffected parents. Following a systematic analysis, 29 of 145 rare protein altering DNMs were classified as possibly causative of the male infertility phenotype. We observed a significant enrichment of Loss-of-Function (LoF) DNMs in LoF-intolerant genes (p-value=1.00x10-5) as well as predicted pathogenic missense DNMs in missense-intolerant genes (p-value=5.01x10-4). One DNM gene identified, RBM5, is an essential regulator of male germ cell pre-mRNA splicing2. In a follow-up study, 5 rare pathogenic missense mutations affecting this gene were observed in a cohort of 2,279 infertile patients, with no such mutations found in a cohort of 5,784 fertile men (p-value=0.009). Our results provide the first evidence for the role of DNMs in severe male infertility and point to many new candidate genes affecting fertility.

genetics↗

bioRxiv2020

New insights into sperm ultrastructure through enhanced scanning electron microscopy

The analysis of spermatozoa morphology is fundamental to understand male fertility and the aetiology of infertility. Traditionally scanning electron microscopy (SEM) has been used to define surface topology. Recently, however, it has become a critical tool for three-dimensionally analyse of internal cellular ultrastructure. Modern SEM provides nanometer-scale resolution, but the meaningfulness of such information is proportional to the quality of the sample preservation. In this study, we demonstrate that sperm quickly and robustly adhere to gold-coated surfaces. Leveraging this property, we developed three step-by-step protocols fulfilling different needs for sperm imaging: chemically fixed monolayers for SEM examination of the external morphology, and two high-pressure freezing-based protocols for fast SEM examination of full cell internal morphology and focused ion-beam SEM (FIB-SEM) tomography. These analyses allow previously unappreciated insights into mouse sperm ultrastructure, including the identification of novel structures within the fibrous sheath and domain-specific interactions between the plasma membrane and exosome-like structures.

cell biology↗

bioRxiv2018

Oxygen plasma focused ion beam scanning electron microscopy for biological samples

Over the past decade, gallium Focused Ion Beam-Scanning Electron Microscopy (FIB-SEM) has been established as a key technology for cellular tomography. The utility of this approach, however, is severely limited both by throughput and the limited selection of compatible sample preparation protocols. Here, we address these limitations and present oxygen plasma FIB (O-PFIB) as a new and versatile tool for cellular FIB-SEM tomography. Oxygen displays superior resin compatibility to other ion beams and produces curtain-free surfaces with minimal polishing. Our novel approach permits more flexible sample preparation and 30% faster data collection when compared to using gallium ion sources. We demonstrate this alternative FIB is applicable to a variety of embedding procedures and biological samples including brain tissue and whole organisms. Finally, we demonstrate the use of O-PFIB to produce targeted FIB-SEM tomograms through fiducial free en-block correlative light and electron microscopy.

cell biology↗