Spastin is a conserved microtubule-severing enzyme mutated in hereditary spastic paraplegia. The role that spastin plays in the cell biology of axon regeneration and degeneration has recently been investigated in Drosophila . We show that the C. elegans spastin ortholog, spas-1 , is expressed in GABA motor neurons, in addition to the known expression in touch receptor neurons (TRNs) and that it is required for axon regeneration in the GABA motor neurons after in vivo laser axotomy. We identified no neuronal developmental defects in the GABA motor neurons and only minor branching variations in the TRNs. However, we show that spas-1 is required for the long-term maintenance of TRN axons in C. elegans , as older spas-1 null C. elegans show a significant increase in specific axonal morphological defects compared with the wild type as identified by confocal microscopy in aged animals. Together, our results suggest that spastin is required for regrowth and maintenance of axons in C. elegans , consistent with pre...

Rigorous, statistically grounded experimental design is central to ethical and effective animal research. Foundational principles for statistically based Design of Experiments (DOE) were established over a century ago by Sir Ronald Fisher. They have since been augmented by modern computational tools that now enable researchers to implement designs that maximize scientific information and benefit while minimizing harms. However, many preclinical investigators are unfamiliar with formal DOE methods. Poorly designed experiments followed by inappropriate statistical analyses contribute to poor reproducibility, translational failure, and unnecessary animal use. This first paper in a three-part series introduces neuroscience researchers to the fundamentals of statistically based experimental design as a substitute for traditional two-group comparisons. Key components of a designed experiment are defined, along with the importance of correctly identifying experimental units to avoid pseudo-replication. Fisher's t...

The progressive ratio (PR) schedule is a popular and well-established tool used to study decision-making and effort across species. In this task, subjects perform an instrumental response to receive a reinforcement, but the ratio requirement increases throughout the behavioral session. A subject's breakpoint, or the point at which the subject is no longer willing to exert the effort required to receive reinforcement, is the main behavioral readout analyzed in the PR schedule. The PR schedule has been used to investigate motivation for various reinforcers (e.g., drugs of abuse) and has been adapted to incorporate aspects of effort-related decision-making (e.g., choice between high- and low-value outcomes). However, there are notable limitations to the utility of the PR as a test of decision-making, including the confounding effects of effort and delay in that higher ratios take longer to complete, the limited behavioral response options (e.g., lever pressing), and breakpoint being a low-dimensional measurem...

Natural environments contain behaviorally relevant information along many stimulus dimensions, each of which sensory systems must encode in order to guide behaviors. For example, the mammalian visual cortex encodes features of visual scenes such as spatial information related to object identity and temporal information about the motion of those objects in space. In order to reliably encode these behaviorally relevant visual features, neural representations should be robust to changes in environmental conditions. Further, information about changes in environmental conditions, such as the luminance changes that occur over the course of a day, is also important for guiding behaviors. In this study, we asked whether mouse primary visual cortex (V1) jointly represents the spatial properties of visual stimuli along with changes in the mean luminance of the visual scene. We find that while V1 neurons, in mice of either sex, encode spatial aspects of visual information in an invariant manner across luminance condi...

Nonrapid eye movement (NREM) sleep is characterized by the interaction of multiple oscillations essential for memory consolidation, alongside a dynamic arrhythmic 1/ f scale-free background that may also contribute to its functions. Recent spectral parametrization methods, such as fitting oscillation and one-and-over-F and irregular resampling auto-spectral analysis, enable the dissociation of rhythmic and arrhythmic components in the spectral domain; however, they do not resolve these processes in the time domain. Instantaneous measures of frequency, amplitude, and phase–amplitude coupling (PAC) are thus still confounded by fluctuations in arrhythmic activity. This limitation represents a pitfall for studies of NREM sleep relying on instantaneous estimates to investigate oscillatory coupling. To address this limitation, we introduce “Rhythms and Background” (RnB), a novel wavelet-based methodology designed to dynamically denoise time series data of arrhythmic interference. This enables the extraction of p...

Dementia-causing diseases, including Alzheimer's disease (AD), are one of the greatest health concerns facing the aging world population. A key feature of AD is excessive accumulation of amyloid-beta, leading to synapse and cell loss in brain structures, such as the hippocampus. This neurodegeneration is preceded by impaired neuron function, notably reduced synaptic inhibition. Metabotropic GABAB receptors (GABABRs) may be modulated by amyloid precursor protein (APP) and are reported to be progressively lost from neuronal membranes of hippocampal pyramidal neurons. However, it remains unknown whether functional GABABR-mediated signaling changes over aging and whether or not pharmacological intervention can prevent receptor loss. In this study, we combine electrophysiological and biochemical analysis of hippocampal neurons in the Amyloid Precursor Protein/Presenilin-1 (APP/PS1) mouse model of AD from acute brain slices and organotypic slice cultures prepared from male and female mice to determine if functio...

Neurexins (Nrxns) are presynaptic cell adhesion molecules essential for synapse development and function. Of the many neurexin isoforms, only β-Nrxns contain the histidine-rich domain (HRD). While the HRD has been implicated in several pathological contexts, its normal physiological role remains unclear. To address this, we used a CRISPR-Cas9 method to generate a new mouse line expressing in-frame truncated Nrxn1β lacking the HRD. We found that HRD deletion did not affect mouse viability, gross brain development, or general behavior of either sex. However, loss of the HRD significantly altered neuroligin-1-dependent excitatory, but not inhibitory, presynaptic differentiation in primary cultured neurons. Moreover, this deletion affected presynaptic short-term plasticity, but not basal synaptic transmission, at hippocampal Schaffer collateral→CA1 synapses. These findings identify the Nrxn1β HRD as a potential contributor to excitatory presynaptic organization and function, providing new insight into the mole...

Blocking is a key statistical method introduced almost a century ago by Ronald Fisher. Blocking controls the effect of “nuisance” variables that are not of direct interest but introduce unwanted variation into the experimental response. Block factors, such as cage, litter, or time, are used to group experimental units into homogeneous subsets. There are two types of block designs: complete and incomplete. In complete block designs every treatment appears in every block. Examples include the Randomized Complete Block Design (RCBD) with a single block factor, and variants such as Latin square and Graeco-Latin square designs with multiple block factors. RCBDs are simple, flexible, and the most widely used. Replicated and nested Latin square designs allow more rigorous control of complex nuisance structures with minimal sample size. Incomplete block designs are extremely useful when practical constraints (e.g., caging density or varying litter sizes) restrict complete treatment replication across all blocks. B...

Behavioral neuroscientists are in the business of linking neuron function to behavior. Historically, single-unit recording has done much of the heavy lifting in this work. Nobel Prize-winning work on the visual system came from experiments such as Hubel and Wiesel (1962), in which 303 cortical neurons were painstakingly recorded across 40 subjects. Single-neuron firing was examined under a wide range of visual conditions. Careful evaluation of single-neuron firing, combined with anatomy and connectivity, uncovered the functional organization of the visual system. “What is my neuron doing?” is one of the most important and powerful questions in behavioral neuroscience. Modern neuroscience has seen a torrent of new tools for monitoring neural activity. Single-unit recording can now track hundreds of neurons per session (Steinmetz et al., 2021; Song et al., 2024). Emerging voltage indicators are providing direct optical readouts of membrane potential in behaving subjects (Hao et al., 2024). Yet no technique ...

Vestibular dysfunction constitutes a major medical concern, and regeneration of hair cells (HCs) is a primary target of gene therapy aimed at restoring vestibular functions. Thus far, therapeutic trials in animal models targeting vestibular loss associated with genetic diseases have yielded variable and partial results, and the functional identity and quantity of HCs required to restore minimal or normal vestibular function remain undefined. Indeed, direct comparisons between structural pathology and quantitative assessments of vestibular dysfunctions are lacking in humans and are rather limited in animal models, representing a significant gap in current knowledge. Here, we present an innovative methodology to bridge the gap between HC integrity and functional vestibular loss in individual mice of either sex. Gradual vestibular deficits were induced through a dose-dependent ototoxic lesion, quantified with canal or utricular-specific vestibulo-ocular reflex tests, and were then correlated in all individual...