Running-induced neurogenesis reduces CA1 perineuronal net density without substantial temporal delay.

Aerobic exercise has many effects on brain function, particularly at the hippocampus. Exercise has been shown to increase the rate of adult neurogenesis within the dentate gyrus and decrease the density of perineuronal nets in area CA1. The relationship between the rate of neurogenesis and the density of perineuronal nets in CA1 is robust; however, these studies only ever examined these effects across longer time scales, with running manipulations of 4 weeks or longer. With such long periods of manipulation, the precise temporal nature of the relationship between running-induced neurogenesis and reduced perineuronal net density in CA1 is unknown. Here, we provided male and female mice with home cage access to running wheels for 0, 1, 2, or 4 weeks and quantified hippocampal neurogenesis and CA1 perineuronal net density. In doing so, we observed a 2-week delay period prior to the increase in neurogenesis, which coincided with the same delay prior to decreased CA1 perineuronal net density. These results highlight the closely linked temporal relationship between running-induced neurogenesis and decreased perineuronal net expression in CA1.

Long-Term Impact of Early-Life Stress on Serotonin Connectivity.

BACKGROUND: Chronic childhood stress is a prominent risk factor for developing affective disorders, yet mechanisms underlying this association remain unclear. Maintenance of optimal serotonin (5-HT) levels during early postnatal development is critical for the maturation of brain circuits. Understanding the long-lasting effects of early-life stress (ELS) on serotonin-modulated brain connectivity is crucial to develop treatments for affective disorders arising from childhood stress. METHODS: Using a mouse model of chronic developmental stress, we determined the long-lasting consequences of ELS on 5-HT circuits and behavior in females and males. Using FosTRAP mice, we cross-correlated regional c-Fos density to determine brain-wide functional connectivity of the raphe nucleus. We next performed in vivo fiber photometry to establish ELS-induced deficits in 5-HT dynamics and optogenetics to stimulate 5-HT release to improve behavior. RESULTS: Adult female and male mice exposed to ELS showed heightened anxiety-like behavior. ELS further enhanced susceptibility to acute stress by disrupting the brain-wide functional connectivity of the raphe nucleus and the activity of 5-HT neuron population, in conjunction with increased orbitofrontal cortex (OFC) activity and disrupted 5-HT release in medial OFC. Optogenetic stimulation of 5-HT terminals in the medial OFC elicited an anxiolytic effect in ELS mice in a sex-dependent manner. CONCLUSIONS: These findings suggest a significant disruption in 5-HT-modulated brain connectivity in response to ELS, with implications for sex-dependent vulnerability. The anxiolytic effect of the raphe-medial OFC circuit stimulation has potential implications for developing targeted stimulation-based treatments for affective disorders that arise from early life adversities.

PAW, a cost-effective and open-source alternative to commercial rodent running wheels.

Voluntary wheel running is a common measure of general activity in many rodent models across neuroscience and physiology. However, current commercial wheel monitoring systems can be cost-prohibitive to many investigators, with many of these systems requiring investments of thousands of dollars. In recent years, several open-source alternatives have been developed, and while these tools are much more cost effective than commercial system, they often lack the flexibility to be applied to a wide variety of projects. Here, we have developed PAW, a 3D Printable Arduino-based Wheel logger. PAW is wireless, fully self-contained, easy to assemble, and all components necessary for its production can be obtained for only $75 CAD. Furthermore, with its compact internal electronics, the 3D printed casing can be easily modified to be used with a wide variety of running wheel designs for a wide variety of rodent species. Data recorded with the PAW system shows circadian patterns of activity which is expected from mice and is consistent with results found in the literature. Altogether, PAW is a flexible, low-cost system that can be beneficial to a broad range of researchers who study rodent models.

Protocol for the integration of fiber photometry and social behavior in rodent models.

Fiber photometry offers insight into cell-type-specific activity underlying social interactions. We provide a protocol for the integration of fiber photometry recordings into the analysis of social behavior in rodent models. This includes considerations during surgery, notes on synchronizing fiber photometry with behavioral recordings, advice on using multi-animal behavioral tracking software, and scripts for the analysis of fiber photometry recordings. For complete details on the use and execution of this protocol, please refer to Dawson et al. (2023).1.

Parvalbumin as a sex-specific target in Alzheimer's disease research - A mini-review.

Alzheimer's disease (AD) is the most common form of dementia, and both the incidence of this disease and its associated cognitive decline disproportionally effect women. While the etiology of AD is unknown, recent work has demonstrated that the balance of excitatory and inhibitory activity across the brain may serve as a strong predictor of cognitive impairments in AD. Across the cortex, the most prominent source of inhibitory signalling is from a class of parvalbumin-expressing interneurons (PV+). In this mini-review, the impacts of sex- and age-related factors on the function of PV+ neurons are examined within the context of vulnerability to AD pathology. These primary factors of influence include changes in brain metabolism, circulating sex hormone levels, and inflammatory response. In addition to positing the increased vulnerability of PV+ neurons to dysfunction in AD, this mini-review highlights the critical importance of presenting sex stratified data in the study of AD.

Hypocretin/orexin neurons encode social discrimination and exhibit a sex-dependent necessity for social interaction.

The hypothalamus plays a crucial role in the modulation of social behavior by encoding internal states. The hypothalamic hypocretin/orexin neurons, initially identified as regulators of sleep and appetite, are important for emotional and motivated behaviors. However, their role in social behavior remains unclear. Using fiber photometry and behavioral analysis, we show here that hypocretin neurons differentially encode social discrimination based on the nature of social encounters. The optogenetic inhibition of hypocretin neuron activity or blocking of hcrt-1 receptors reduces the amount of time mice are engaged in social interaction in males but not in females. Reduced hcrt-1 receptor signaling during social interaction is associated with altered activity in the insular cortex and ventral tegmental area in males. Our data implicating hypocretin neurons as sexually dimorphic regulators within social networks have significant implications for the treatment of neuropsychiatric diseases with social dysfunction, particularly considering varying prevalence among sexes.

Low intensity repetitive transcranial magnetic stimulation modulates brain-wide functional connectivity to promote anti-correlated c-Fos expression.

Repetitive transcranial magnetic stimulation (rTMS) induces action potentials to induce plastic changes in the brain with increasing evidence for the therapeutic importance of brain-wide functional network effects of rTMS; however, the influence of sub-action potential threshold (low-intensity; LI-) rTMS on neuronal activity is largely unknown. We investigated whether LI-rTMS modulates neuronal activity and functional connectivity and also specifically assessed modulation of parvalbumin interneuron activity. We conducted a brain-wide analysis of c-Fos, a marker for neuronal activity, in mice that received LI-rTMS to visual cortex. Mice received single or multiple sessions of excitatory 10 Hz LI-rTMS with custom rodent coils or were sham controls. We assessed changes to c-Fos positive cell densities and c-Fos/parvalbumin co-expression. Peak c-Fos expression corresponded with activity during rTMS. We also assessed functional connectivity changes using brain-wide c-Fos-based network analysis. LI-rTMS modulated c-Fos expression in cortical and subcortical regions. c-Fos density changes were most prevalent with acute stimulation, however chronic stimulation decreased parvalbumin interneuron activity, most prominently in the amygdala and striatum. LI-rTMS also increased anti-correlated functional connectivity, with the most prominent effects also in the amygdala and striatum following chronic stimulation. LI-rTMS induces changes in c-Fos expression that suggest modulation of neuronal activity and functional connectivity throughout the brain. Our results suggest that LI-rTMS promotes anticorrelated functional connectivity, possibly due to decreased parvalbumin interneuron activation induced by chronic stimulation. These changes may underpin therapeutic rTMS effects, therefore modulation of subcortical activity supports rTMS for treatment of disorders involving subcortical dysregulation.

Brain-wide neuronal activation and functional connectivity are modulated by prior exposure to repetitive learning episodes.

Memory storage and retrieval are shaped by past experiences. Prior learning and memory episodes have numerous impacts on brain structure from micro to macroscale. Previous experience with specific forms of learning increases the efficiency of future learning. It is less clear whether such practice effects on one type of memory might also have transferable effects to other forms of memory. Different forms of learning and memory rely on different brain-wide networks but there are many points of overlap in these networks. Enhanced structural or functional connectivity caused by one type of learning may be transferable to another type of learning due to overlap in underlying memory networks. Here, we investigated the impact of prior chronic spatial training on the task-specific functional connectivity related to subsequent contextual fear memory recall in mice. Our results show that mice exposed to prior spatial training exhibited decreased brain-wide activation compared to control mice during the retrieval of a context fear memory. With respect to functional connectivity, we observed changes in several network measures, notably an increase in global efficiency. Interestingly, we also observed an increase in network resilience based on simulated targeted node deletion. Overall, this study suggests that chronic learning has transferable effects on the functional connectivity networks of other types of learning and memory. The generalized enhancements in network efficiency and resilience suggest that learning itself may protect brain networks against deterioration.

New neurons in old brains: implications of age in the analysis of neurogenesis in post-mortem tissue.

Adult neurogenesis, the proliferation and integration of newly generated neurons, has been observed in the adult mammalian hippocampus of many species. Numerous studies have also found adult neurogenesis in the human hippocampus, but several recent high-profile studies have suggested that this process is considerably reduced in humans, occurring in children but not in adults. In comparison, rodent studies also show age-related decline but a greater degree of proliferation of new neurons in adult animals. These differences may represent biological species differences or could alternatively be explained by methodological differences in tissue handling and fixation. Here, we examine whether differences in the post-mortem interval between death and tissue fixation might impact subsequent detection of adult neurogenesis due to increased tissue degradation. Because there are fewer new neurons present in older subjects to begin with we hypothesized that, subject age might interact significantly with post-mortem interval in the detection of adult neurogenesis. We analyzed neurogenesis in the hippocampus of rats that were either perfusion-fixed or the brains extracted and immersion-fixed at various post-mortem intervals. We observed an interaction between animal age and the time delay between death and tissue fixation. While similar levels of neurogenesis were observed in young rats regardless of fixation, older rats had significantly fewer labeled neurons when fixation was not immediate. Furthermore, the morphological detail of the labeled neurons was significantly reduced in the delayed fixation conditions at all ages. This study highlights critical concerns that must be considered when using post-mortem tissue to quantify adult neurogenesis.

Neurogenesis mediated plasticity is associated with reduced neuronal activity in CA1 during context fear memory retrieval.

Postnatal hippocampal neurogenesis has been demonstrated to affect learning and memory in numerous ways. Several studies have now demonstrated that increased neurogenesis can induce forgetting of memories acquired prior to the manipulation of neurogenesis and, as a result of this forgetting can also facilitate new learning. However, the mechanisms mediating neurogenesis-induced forgetting are not well understood. Here, we used a subregion-based analysis of the immediate early gene c-Fos as well as in vivo fiber photometry to determine changes in activity corresponding with neurogenesis induced forgetting. We found that increasing neurogenesis led to reduced CA1 activity during context memory retrieval. We also demonstrate here that perineuronal net expression in areas CA1 is bidirectionally altered by the levels or activity of postnatally generated neurons in the dentate gyrus. These results suggest that neurogenesis may induce forgetting by disrupting perineuronal nets in CA1 which may otherwise protect memories from degradation.

FASTMAP: Open-Source Flexible Atlas Segmentation Tool for Multi-Area Processing of Biological Images.

To better understand complex systems, such as the brain, studying the interactions between multiple brain regions is imperative. Such experiments often require delineation of multiple brain regions on microscopic images based on preexisting brain atlases. Experiments examining the relationships of multiple regions across the brain have traditionally relied on manual plotting of regions. This process is very intensive and becomes untenable with a large number of regions of interest (ROIs). To reduce the amount of time required to process multi-region datasets, several tools for atlas registration have been developed; however, these tools are often inflexible to tissue type, only supportive of a limited number of atlases and orientation, require considerable computational expertise, or are only compatible with certain types of microscopy. To address the need for a simple yet extensible atlas registration tool, we have developed FASTMAP, a Flexible Atlas Segmentation Tool for Multi-Area Processing. We demonstrate its ability to register images efficiently and flexibly to custom mouse brain atlas plates, to detect differences in the regional numbers of labels of interest, and to conduct densitometry analyses. This open-source and user-friendly tool will facilitate the atlas registration of diverse tissue types, unconventional atlas organizations, and a variety of tissue preparations.

Network Neuroscience Untethered: Brain-Wide Immediate Early Gene Expression for the Analysis of Functional Connectivity in Freely Behaving Animals.

Studying how spatially discrete neuroanatomical regions across the brain interact is critical to advancing our understanding of the brain. Traditional neuroimaging techniques have led to many important discoveries about the nature of these interactions, termed functional connectivity. However, in animal models these traditional neuroimaging techniques have generally been limited to anesthetized or head-fixed setups or examination of small subsets of neuroanatomical regions. Using the brain-wide expression density of immediate early genes (IEG), we can assess brain-wide functional connectivity underlying a wide variety of behavioural tasks in freely behaving animal models. Here, we provide an overview of the necessary steps required to perform IEG-based analyses of functional connectivity. We also outline important considerations when designing such experiments and demonstrate the implications of these considerations using an IEG-based network dataset generated for the purpose of this review.

Adult neurogenesis mediates forgetting of multiple types of memory in the rat.

The formation and retention of hippocampus-dependent memories is impacted by neurogenesis, a process that involves the production of new neurons in the dentate gyrus of the hippocampus. Recent studies demonstrate that increasing neurogenesis after memory formation induces forgetting of previously acquired memories. Neurogenesis-induced forgetting was originally demonstrated in mice, but a recent report suggests that the same effect may be absent in rats. Although a general species difference is possible, other potential explanations for these incongruent findings are that memories which are more strongly reinforced become resilient to forgetting or that perhaps only certain types of memories are affected. Here, we investigated whether neurogenesis-induced forgetting occurs in rats using several hippocampus-dependent tasks including contextual fear conditioning (CFC), the Morris Water Task (MWT), and touchscreen paired associates learning (PAL). Neurogenesis was increased following training using voluntary exercise for 4 weeks before recall of the previous memory was assessed. We show that voluntary running causes forgetting of context fear memories in a neurogenesis-dependent manner, and that neurogenesis-induced forgetting is present in rats across behavioral tasks despite differences in complexity or reliance on spatial, context, or object memories. In addition, we asked whether stronger memories are less susceptible to forgetting by varying the strength of training. Even with a very strong training protocol in the CFC task, we still observed enhanced forgetting related to increased neurogenesis. These results suggest that forgetting due to neurogenesis is a conserved mechanism that aids in the clearance of memories.

Standardised ginseng extract G115® potentiates the antidepressant-like properties of fluoxetine in the forced swim test.

OBJECTIVE: Ginsenosides, biologically active components of the root of Panax ginseng, have been reported to have therapeutic benefits in a number of disease states including psychiatric conditions such as major depressive disorder. Our objective was to determine if a standardised commercial ginseng extract, G115®, could reduce the signs of behavioural despair commonly observed in animal models of depression either alone or in combination with the selective serotonin reuptake inhibitor (SSRI) fluoxetine. METHODS: Male Sprague-Dawley (SD) rats (N = 51) were divided into four groups: vehicle control, G115® ginseng root extract, fluoxetine and fluoxetine plus G115®. Rats were trained to voluntarily consume treatments twice daily for 14 days and were then tested in an open field (OF), elevated plus maze (EPM) and forced swim test (FST). Post-mortem hippocampal and prefrontal cortex tissue was analysed for expression of brain-derived neurotrophic factor (BDNF) and tropomyosin receptor kinase B (TrkB) by western blot. RESULTS: One-way Analysis of Variance revealed no significant group differences in the OF or plus-maze performance on any variable examined. In the FST, fluoxetine significantly reduced immobility time and increased latency to immobility. The effects of fluoxetine were further significantly potentiated by co-administration of G115®. Post-mortem tissue analysis revealed significant group differences in BDNF expression in the left hippocampus and left prefrontal cortex without any accompanying changes in TrkB expression. CONCLUSIONS: We conclude that oral G115® significantly potentiates the antidepressant-like effect of fluoxetine in the FST in the absence of potentially confounding effects on locomotion and anxiety.

Disrupted Neurogenesis in Germ-Free Mice: Effects of Age and Sex.

The gut microbiome has profound effects on development and function of the nervous system. Recent evidence indicates that disruption of the gut microbiome leads to altered hippocampal neurogenesis. Here, we examined whether the effects of gut microbiome disruption on neurogenesis are age-dependent, given that both neurogenesis and the microbiome show age-related changes. Additionally, we examined memory induced functional connectivity of hippocampal networks. Control and germ-free mice at three different ages (4, 8, and 12 weeks) were trained in contextual fear-conditioning, then subsequently tested the following day. Hippocampal neurogenesis, quantified via BrdU and doublecortin, exhibited age-dependent changes relative to controls, with the established age-dependent decrease in neurogenesis being delayed in germ-free mice. Moreover, we found sex-dependent effects of germ-free status on neurogenesis, with 4 week old male germ-free mice having decreased neurogenesis and 8 week old female germ-free mice having increased neurogenesis. To assess systems-level consequences of disrupted neurogenesis, we assessed functional connectivity of hippocampal networks by inducing c-Fos expression with contextual memory retrieval and applying a previously described network analysis. Our results indicate impaired connectivity of the dentate gyrus in germ-free mice in a pattern highly correlated with adult neurogenesis. In control but not germ-free mice, functional connectivity became more refined with age, indicating that age dependent network refinement is disrupted in germ-free mice. Overall, the results show that disruption of the gut microbiome affects hippocampal neurogenesis in an age- and sex-dependent manner and that these changes are also related to changes in the dentate gyrus functional network.