Sensorimotor circuits of the lumbosacral spinal cord are required for lower urinary tract (LUT) regulation as well as being engaged in pelvic pain states. To date, no molecular markers have been identified to enable specific visualization of LUT afferents, which are embedded within spinal cord segments that also subserve somatic functions. Moreover, previous studies have not fully investigated the patterning within or across spinal segments, compared afferent innervation of the bladder and urethra, or explored possible structural sex differences in these pathways. We have addressed these questions in adult Sprague Dawley rats, using intramural microinjection of the tract tracer, B subunit of cholera toxin (CTB). Afferent distribution was analyzed within individual sections and 3D reconstructions from sections across four spinal cord segments (L5-S2), and in cleared intact spinal cord viewed with light sheet microscopy. Simultaneous mapping of preganglionic neurons showed their location throughout S1 but re...

The activity-dependent expression of immediate-early genes (IEGs) has been utilized to label memory traces. However, their roles in engram specification are incompletely understood. Outstanding questions remain as to whether expression of IEGs can interplay with network properties such as functional connectivity and also if neurons expressing different IEGs are functionally distinct. In order to connect IEG expression at the cellular level with changes in functional connectivity, we investigated the expression of 2 IEGs, Arc and c-Fos, in cultured hippocampal neurons. Primary neuronal cultures were treated with a chemical cocktail [4-aminopyridine (4AP), bicuculline (Bic), and forskolin] to increase neuronal activity, IEG expression, and induce chemical long-term potentiation (LTP). Neuronal firing is assayed by intracellular calcium imaging using GCaMP6m and expression of IEGs is assessed by immunofluorescence staining. We noted an emergent network property of refinement in network activity, characterized...

Water restriction is commonly used to motivate rodents to perform behavioral tasks; however, its effects on hydration and stress hormone levels are unknown. Here, we report daily body weight and bi-weekly packed red blood cell volume and corticosterone (CORT) in adult male rats across 80 days for three commonly used water restriction schedules. We also assessed renal adaptation to water restriction using postmortem histologic evaluation of renal medulla. A control group received ad libitum water. After one week of water restriction, rats on all restriction schedules resumed similar levels of growth relative to the control group. Normal hydration was observed, and water restriction did not drive renal adaptation. An intermittent restriction schedule was associated with an increase in CORT relative to the control group. However, intermittent restriction evokes a stress response which could affect behavioral and neurobiological results. Our results also suggest that stable motivation in behavioral tasks may o...

Vision loss from diseases of the outer retina, such as age-related macular degeneration, is among the leading causes of irreversible blindness in the world today. The goal of retinal prosthetics is to replace the photo-sensing function of photoreceptors lost in these diseases with optoelectronic hardware to electrically stimulate patterns of retinal activity corresponding to vision. To enable high-resolution retinal prosthetics, the scale of stimulating electrodes must be significantly decreased from current designs; however, this reduces the amount of stimulating current that can be delivered. The efficacy of subretinal stimulation at electrode sizes suitable for high visual acuity retinal prosthesis are not well understood, particularly within the safe charge injection limits of electrode materials. Here, we measure retinal ganglion cell (RGC) responses in a mouse model of blindness to evaluate the stimulation efficacy of 10, 20, and 30 μm diameter iridium oxide electrodes within the electrode charge inj...

Hyperglycemia is a comorbidity in 60–80% of stroke patients; nevertheless, neuroprotective drugs like NMDA receptor (NMDAR) antagonists are typically assessed in normoglycemic animals at the preclinical stage before they are approved to enter clinical trials. Interestingly, as a possible explanation for the translational failure of NMDAR antagonists, it was recently reported that stroke occurring during nighttime causes smaller infarctions in rodents and therefore has a smaller window for neuroprotection. To investigate why stroke occurring during different circadian phases confers a difference in severity, we reanalyzed the published source data and found that some mice that were used in the daytime have higher blood glucose than mice that were used in the nighttime. We then repeated the experiments but found no difference in blood glucose concentration or infarct volume regardless of the circadian phase during which stroke occurs. On the other hand, induction of hyperglycemia by glucose injection reprodu...

The cochaperone FKBP51, encoded by the Fkbp5 gene, has been identified as central risk factor for anxiety-related disorders and stress system dysregulation. In the brain, the oval bed nucleus of the stria terminalis (ovBNST) has been implicated in stress-induced anxiety. However, the role of Fkbp5 in the ovBNST and its impact on anxiety-like behavior have remained unknown. Here, we show in mice that Fkbp5 in the ovBNST is reactive to acute stress and coexpressed with the stress-regulated neuropeptides Tac2 and Crh . Subsequently, results obtained from viral-mediated manipulation indicate that Fkbp5 overexpression (OE) in the ovBNST results in an anxiolytic-like tendency regarding behavior and endocrinology, whereas a Fkbp5 knock-out (KO) exposed a clear anxiogenic phenotype, indicating that native ovBNST expression and regulation is necessary for normal anxiety-related behavior. Notably, our data suggests that a stress-induced increase of Fkbp5 in the ovBNST may in fact have a protective role, leading to a...

Multisensory plasticity enables our senses to dynamically adapt to each other and the external environment, a fundamental operation that our brain performs continuously. We searched for neural correlates of adult multisensory plasticity in the dorsal medial superior temporal area (MSTd) and the ventral intraparietal area (VIP) in two male rhesus macaques using a paradigm of supervised calibration. We report little plasticity in neural responses in the relatively low-level multisensory cortical area MSTd. In contrast, neural correlates of plasticity are found in higher level multisensory VIP, an area with strong decision-related activity. Accordingly, we observed systematic shifts of VIP tuning curves, which were reflected in the choice-related component of the population response. This is the first demonstration of neuronal calibration, together with behavioral calibration, in single sessions. These results lay the foundation for understanding multisensory neural plasticity, applicable broadly to maintaini...

Tracking and quantifying the abundance and location of cells in the developing brain is essential in neuroscience research, enabling a greater understanding of mechanisms underlying nervous system morphogenesis. Widely used experimental methods to quantify cells labeled with fluorescent markers—such as immunohistochemistry, in situ hybridization, and expression of transgenes via stable lines or transient in utero electroporations—depend upon accurate and consistent quantification of images. Current methods to quantify fluorescently-labeled cells rely on labor-intensive manual counting approaches, such as the Fiji plugin Cell Counter , which requires custom macros to enable higher-throughput analyses. Here, we present RapID Cell Counter, a semi-automated cell-counting tool with an easy-to-implement graphical user interface, that facilitates quick and consistent quantifications of cell density within user-defined boundaries that can be divided into equally-partitioned segments. Compared to the standard manua...

With the wide adoption of genomic sequencing in children having seizures, an increasing number of SCN2A genetic variants have been revealed as genetic causes of epilepsy. Voltage-gated sodium channel Nav1.2, encoded by gene SCN2A , is predominantly expressed in the pyramidal excitatory neurons and supports action potential firing. One recurrent SCN2A genetic variant is L1342P, which was identified in multiple patients with epileptic encephalopathy and intractable seizures. However, the mechanism underlying L1342P-mediated seizures and the pharmacogenetics of this variant in human neurons remain unknown. To understand the core phenotypes of the L1342P variant in human neurons, we took advantage of a reference human induced pluripotent stem cell (hiPSC) line from a male donor, in which L1342P was introduced by CRISPR/Cas9 mediated genome editing. Using patch-clamping and micro-electrode array (MEA) recordings, we revealed that cortical neurons derived from hiPSCs carrying heterozygous L1342P variant have sig...

Despite being canonically considered a motor control structure, the cerebellum is increasingly recognized for important roles in processes beyond this traditional framework, including seizure suppression. Excitatory fastigial neurons project to a large number of downstream targets, and it is unclear if this broad targeting underlies seizure suppression, or if a specific output may be sufficient. To address this question, we used the intrahippocampal kainic acid mouse model of temporal lobe epilepsy, male and female animals, and a dual-virus approach to selectively label and manipulate fastigial outputs. We examined fastigial neurons projecting to the superior colliculus, medullary reticular formation, and central lateral nucleus of the thalamus, and found that these comprise largely non-overlapping populations of neurons which send collaterals to unique sets of additional, somewhat overlapping, thalamic and brainstem regions. We found that neither optogenetic stimulation of superior colliculus nor reticula...