Rewarded Maze Training Increases Approach Behavior in Rats Through Neurogenesis-Dependent Growth of Ventral Hippocampus-Prelimbic Circuits.

Background: Learning complex navigation routes increases hippocampal volume in humans, but it is not clear whether this growth impacts behaviors outside the learning situation or what cellular mechanisms are involved. Methods: We trained rats with pharmacogenetic suppression of adult neurogenesis and littermate controls in 3 mazes over 3 weeks and tested novelty approach behavior several days after maze exposure. We then measured hippocampus and prelimbic cortex volumes using magnetic resonance imaging and assessed neuronal and astrocyte morphology. Finally, we investigated the activation and behavioral role of the ventral CA1 (vCA1)-to-prelimbic pathway using immediate-early genes and DREADDs (designer receptors exclusively activated by designer drugs). Results: Maze training led to volume increase of both the vCA1 region of the hippocampus and the prelimbic region of the neocortex compared with rats that followed fixed paths. Growth was also apparent in individual neurons and astrocytes in these 2 regions, and behavioral testing showed increased novelty approach in maze-trained rats in 2 different tests. Suppressing adult neurogenesis prevented the effects on structure and approach behavior after maze training without affecting maze learning itself. The vCA1 neurons projecting to the prelimbic area were more activated by novelty in maze-trained animals, and suppression of this pathway decreased approach behavior. Conclusions: Rewarded navigational learning experiences induce volumetric and morphologic growth in the vCA1 and prelimbic cortex and enhance activation of the circuit connecting these 2 regions. Both the structural and behavioral effects of maze training require ongoing adult neurogenesis, suggesting a role for new neurons in experience-driven increases in novelty exploration.

Inhibition of Hippocampal Neurogenesis Starting in Adolescence Increases Anxiodepressive Behaviors Amid Stress.

Stressors during the adolescent period can affect development of the brain and have long-lasting impacts on behavior. Specifically, adolescent stress impairs hippocampal neurogenesis and can increase risk for anxiety, depression, and a dysregulated stress response in adulthood. In order to model the functional effects of reduced hippocampal neurogenesis during adolescence, a transgenic neurogenesis ablation rat model was used to suppress neurogenesis during the adolescent period and test anxiodepressive behaviors and stress physiology during adulthood. Wildtype and transgenic (TK) rats were given valganciclovir during the first two weeks of adolescence (4-6 weeks old) to knock down neurogenesis in TK rats. Starting in young adulthood (13 weeks old), blood was sampled for corticosterone at several time points following acute restraint stress to measure negative feedback of the stress response, and rats were tested on a battery of anxiodepressive tests at baseline and following acute restraint stress. Although TK rats had large reductions in both cell proliferation during adolescence, as measured by bromodeoxyuridine (BrdU), and ongoing neurogenesis in adulthood (by doublecortin), resulting in decreased volume of the dentate gyrus, negative feedback of the stress response following acute restraint was similar across all rats. Despite similar stress responses, TK rats showed higher anxiety-like behavior at baseline. In addition, only TK rats had increased depressive-like behavior when tested after acute stress. Together, these results suggest that long-term neurogenesis ablation starting in adolescence produces hippocampal atrophy and increases behavioral caution and despair amid stressful environments.

Using Zebrafish Embryos to Study Pharmacological Effects on Neural Development in Hands-On Neurobiology Laboratory Activities.

Undergraduate neurobiology courses cover neural development as a major theme but there are few labs to provide hands-on experience with these topics. Here we share a 3-week set of lab activities using zebrafish embryos that allow students to see the direct effect of drug exposure on physical and emotional development. In these labs, student expose new embryos (Lab 1) to the environmental toxin lithium chloride, which inhibits anterior development and produces an eyeless phenotype in fixed larvae (Lab 2), and to psychiatric medications fluoxetine and quetiapine, which alter anxiety-like behavior measured live in grown juveniles (Lab 3). Lab worksheets ask students to investigate the signaling pathways affected by these drugs and how they might affect neural development in different ways. Student opinion surveys suggest these lab activities were successful in both providing hands-on work with zebrafish as a model organism for neural development and better understanding of how drugs can impact development of the nervous system.

A Runner's High for New Neurons? Potential Role for Endorphins in Exercise Effects on Adult Neurogenesis.

Physical exercise has wide-ranging benefits to cognitive functioning and mental state, effects very closely resembling enhancements to hippocampal functioning. Hippocampal neurogenesis has been implicated in many of these mental benefits of exercise. However, precise mechanisms behind these effects are not well known. Released peripherally during exercise, beta-endorphins are an intriguing candidate for moderating increases in neurogenesis and the related behavioral benefits of exercise. Although historically ignored due to their peripheral release and status as a peptide hormone, this review highlights reasons for further exploring beta-endorphin as a key mediator of hippocampal neurogenesis. This includes possible routes for beta-endorphin signaling into the hippocampus during exercise, direct effects of beta-endorphin on cell proliferation and neurogenesis, and behavioral effects of manipulating endogenous opioid signaling. Together, beta-endorphin appears to be a promising mechanism for understanding the specific ways that exercise promotes adult neurogenesis specifically and brain health broadly.

High-fat diet and acute stress have different effects on object preference tests in rats during adolescence and adulthood.

Meals of high-fat diet (HFD) during adolescence produce stronger impairments to memory during adolescence than adulthood, however recovery of memory from adolescent HFD is underexplored. In addition, many tests of rodent memory are confounded by aversive or food-based stimuli, making it difficult to determine baseline memory processing affected by HFD. Thus, we utilized three cohorts of rats (adolescent HFD, adult HFD, and adolescent HFD with recovery) to explore the effects of HFD at different ages on two traditional tests of memory based strictly on object exploration, novel object recognition and novel object location tests. To isolate stress as a variable, rats were tested either at baseline or with cold water swim occurring directly after object acquisition. Results show that preference for novel objects is impaired by stress across all groups, but HFD alone only impairs preference for novel objects during adolescence, although this recovers after switching to a control diet. Additionally, preference for an object in a new location is impaired by HFD in all age groups and fails to recover following diet change. Together the data suggest that stress and HFD differentially affect object preference, based on test type, except during the adolescent period. Because these tests are traditionally interpreted as memory processes dependent on two distinct brain regions, the hippocampus and perirhinal cortex, these results support that stress and HFD affect the hippocampus and perirhinal cortex differently. The data affirm that while perirhinal cortex-dependent behavior recovers, the adolescent period is susceptible to long-lasting dysfunctions of hippocampal behavior by HFD.

Adult neurogenesis alters response to an aversive distractor in a labyrinth maze without affecting spatial learning or memory.

Adult neurogenesis has been implicated in learning and memory of complex spatial environments. However, new neurons also play a role in nonmnemonic behavior, including the stress response and attention shifting. Many commonly used spatial tasks are very simple, and unsuitable for detecting neurogenesis effects, or are aversively motivated, making it difficult to dissociate effects on spatial learning and memory from effects on stress. We have therefore created a novel complex spatial environment, the flex maze, to enable reward-mediated testing of spatial learning in a flexibly configurable labyrinth. Using a pharmacogenetic method to completely inhibit neurogenesis in adulthood, we found that rats lacking new neurons (TK rats) and wild type controls completed and remembered most mazes equally well. However, control rats were slower to complete peppermint-scented mazes than other mazes, while neurogenesis-deficient rats showed no effect of mint on maze behavior, completing these mazes significantly faster than control rats. Additional testing found that wild type and TK rats showed similar detection of, avoidance of, and glucocorticoid response to the mint odor. These results suggest that spatial learning and memory in a labyrinth task is unaffected by the loss of new neurons, but that these cells affect the ability of an aversive stimulus to distract rats from completing the maze.

New neurons restore structural and behavioral abnormalities in a rat model of PTSD.

Post-traumatic stress disorder (PTSD) has been associated with anxiety, memory impairments, enhanced fear, and hippocampal volume loss, although the relationship between these changes remain unknown. Single-prolonged stress (SPS) is a model for PTSD combining three forms of stress (restraint, swim, and anesthesia) in a single session that results in prolonged behavioral effects. Using pharmacogenetic ablation of adult neurogenesis in rats, we investigated the role of new neurons in the hippocampus in the long-lasting structural and behavioral effects of SPS. Two weeks after SPS, stressed rats displayed increased anxiety-like behavior and decreased preference for objects in novel locations regardless of the presence or absence of new neurons. Chronic stress produced by daily restraint for 2 or 6 hr produced similar behavioral effects that were also independent of ongoing neurogenesis. At a longer recovery time point, 1 month after SPS, rats with intact neurogenesis had normalized, showing control levels of anxiety-like behavior. However, GFAP-TK rats, which lacked new neurons, continued to show elevated anxiety-like behavior and enhanced serum corticosterone response to anxiogenic experience. Volume loss in ventral CA1 region of the hippocampus paralleled increases in anxiety-like behavior, occurring in all rats exposed to SPS at the early time point and only rats lacking adult neurogenesis at the later time point. In chronic stress experiments, volume loss occurred broadly throughout the dentate gyrus and CA1 after 6-hr daily stress but was not apparent in any hippocampal subregion after 2-hr daily stress. No effect of SPS was seen on cell proliferation in the dentate gyrus, but the survival of young neurons born a week after stress was decreased. Together, these data suggest that new neurons are important for recovery of normal behavior and hippocampal structure following a strong acute stress and point to the ventral CA1 region as a potential key mediator of stress-induced anxiety-like behavior.

Behavioral and structural adaptations to stress.

Unpredictable aversive experiences, or stressors, lead to changes in depression- and anxiety-related behavior and to changes in hippocampal structure including decreases in adult neurogenesis, granule cell and pyramidal cell dendritic morphology, and volume. Here we review the relationship between these behavioral and structural changes and discuss the possibility that these changes may be largely adaptive. Specifically, we suggest that new neurons in the dentate gyrus enhance behavioral adaptability to changes in the environment, biasing behavior in novel situations based on previous experience with stress. Conversely, atrophy-like changes in the hippocampus and decreased adult neurogenesis following chronic stress may serve to limit stress responses and stabilize behavior during chronic stress.

Magnetic resonance imaging of odorant activity-dependent migration of neural precursor cells and olfactory bulb growth.

Neural progenitors or neuroblasts are produced by precursor cells in the subventricular zone (SVZ) and migrate along the rostral migratory stream (RMS) to the olfactory bulbs (OB) throughout life. In the OB, these adult born neurons either die or replace existing olfactory interneurons, playing a critical role in the stabilization of OB circuitry. Although several aspects of the addition of new neurons into the OB have been studied, it is unclear whether long-distance activity from the OB can regulate the influx of migrating neuroblasts along the RMS. In this study, iron oxide-assisted MRI was used to track the migration of neuroblasts in combination with reversible naris occlusion to manipulate odorant-induced activity. It was found that decreasing olfactory activity led to a decrease in the rate of neuroblast migration along the RMS. Removal of the naris occlusion led to an increase in migratory rate back to control levels, indicating that olfactory activity has regulatory function on neuroblast migration in the RMS. Blocking odorant activity also led to an arrest in OB growth and re-opening the block led to a rapid re-growth returning the bulb size to control levels. Furthermore, pharmacogenetic elimination of the neuroblasts demonstrated that they were required for re-growth of the bulb following sensory deprivation. Together, these results show that sensory activity, neural migration and OB growth are tightly coupled in an interdependent manner.

Stress and Loss of Adult Neurogenesis Differentially Reduce Hippocampal Volume.

BACKGROUND: Hippocampal volume loss is a hallmark of clinical depression. Chronic stress produces volume loss in the hippocampus in humans and atrophy of CA3 pyramidal cells and suppression of adult neurogenesis in rodents. METHODS: To investigate the relationship between decreased adult neurogenesis and stress-induced changes in hippocampal structure and volume, we compared the effects of chronic unpredictable restraint stress and inhibition of neurogenesis in a rat pharmacogenetic model. RESULTS: Chronic unpredictable restraint stress over 4 weeks decreased total hippocampal volume, reflecting loss of volume in all hippocampal subfields and in both dorsal and ventral hippocampus. In contrast, complete inhibition of adult neurogenesis for 4 weeks led to volume reduction only in the dentate gyrus. With prolonged inhibition of neurogenesis for 8 or 16 weeks, volume loss spread to the CA3 region, but not CA1. Combining stress and inhibition of adult neurogenesis did not have additive effects on the magnitude of volume loss but did produce a volume reduction throughout the hippocampus. One month of chronic unpredictable restraint stress and inhibition of adult neurogenesis led to atrophy of pyramidal cell apical dendrites in dorsal CA3 and to neuronal reorganization in ventral CA3. Stress also significantly affected granule cell dendrites. CONCLUSIONS: The findings suggest that adult neurogenesis is required to maintain hippocampal volume but is not responsible for stress-induced volume loss.

Ongoing neurogenesis in the adult dentate gyrus mediates behavioral responses to ambiguous threat cues.

Fear learning is highly adaptive if utilized in appropriate situations but can lead to generalized anxiety if applied too widely. A role of predictive cues in inhibiting fear generalization has been suggested by stress and fear learning studies, but the effects of partially predictive cues (ambiguous cues) and the neuronal populations responsible for linking the predictive ability of cues and generalization of fear responses are unknown. Here, we show that inhibition of adult neurogenesis in the mouse dentate gyrus decreases hippocampal network activation and reduces defensive behavior to ambiguous threat cues but has neither of these effects if the same negative experience is reliably predicted. Additionally, we find that this ambiguity related to negative events determines their effect on fear generalization, that is, how the events affect future behavior under novel conditions. Both new neurons and glucocorticoid hormones are required for the enhancement of fear generalization following an unpredictably cued threat. Thus, adult neurogenesis plays a central role in the adaptive changes resulting from experience involving unpredictable or ambiguous threat cues, optimizing behavior in novel and uncertain situations.

Anxiolytic Actions of Exercise in Absence of New Neurons.

Physical exercise reduces anxiety-like behavior in adult mice. The specific mechanisms that mediate this anxiolytic effect are unclear, but adult neurogenesis in the dentate gyrus has been implicated because it is robustly increased by running and has been linked to anxiodepressive-like behavior. We therefore tested the effects of long-term wheel running on anxiety-like behavior in GFAP-TK (TK) mice, a transgenic strain with complete ablation of adult neurogenesis. Five weeks of running reduced anxiety-like behavior equally in both TK mice and wild type (WT) control mice on two tests, elevated plus-maze and novelty-suppressed feeding. WT and TK mice also had similar patterns of c-fos expression in the hippocampus following anxiety testing. Following testing on the elevated plus-maze, running reduced c-fos expression in the dorsal dentate gyrus and CA3 in both WT and TK mice. Following testing on novelty-suppressed feeding, running reduced c-fos expression throughout the dentate gyrus and CA3 in both WT and TK mice. Interestingly, following testing on a less anxiogenic version of novelty-suppressed feeding, running reduced c-fos expression only in the dorsal dentate gyrus in both WT and TK mice, supporting earlier suggestions that the dorsal hippocampus is less involved in emotional behavior than the ventral region. These results suggest that although running increases adult neurogenesis, new neurons are not involved in the decreased anxiety-like behavior or hippocampal activation produced by running. © 2016 Wiley Periodicals, Inc.

Lasting Adaptations in Social Behavior Produced by Social Disruption and Inhibition of Adult Neurogenesis.

UNLABELLED: Research on social instability has focused on its detrimental consequences, but most people are resilient and respond by invoking various coping strategies. To investigate cellular processes underlying such strategies, a dominance hierarchy of rats was formed and then destabilized. Regardless of social position, rats from disrupted hierarchies had fewer new neurons in the hippocampus compared with rats from control cages and those from stable hierarchies. Social disruption produced a preference for familiar over novel conspecifics, a change that did not involve global memory impairments or increased anxiety. Using the neuropeptide oxytocin as a tool to increase neurogenesis in the hippocampus of disrupted rats restored preference for novel conspecifics to predisruption levels. Conversely, reducing the number of new neurons by limited inhibition of adult neurogenesis in naive transgenic GFAP-thymidine kinase rats resulted in social behavior similar to disrupted rats. Together, these results provide novel mechanistic evidence that social disruption shapes behavior in a potentially adaptive way, possibly by reducing adult neurogenesis in the hippocampus. SIGNIFICANCE STATEMENT: To investigate cellular processes underlying adaptation to social instability, a dominance hierarchy of rats was formed and then destabilized. Regardless of social position, rats from disrupted hierarchies had fewer new neurons in the hippocampus compared with rats from control cages and those from stable hierarchies. Unexpectedly, these changes were accompanied by changes in social strategies without evidence of impairments in cognition or anxiety regulation. Restoring adult neurogenesis in disrupted rats using oxytocin and conditionally suppressing the production of new neurons in socially naive GFAP-thymidine kinase rats showed that loss of 6-week-old neurons may be responsible for adaptive changes in social behavior.

New Hippocampal Neurons Mature Rapidly in Response to Ketamine But Are Not Required for Its Acute Antidepressant Effects on Neophagia in Rats.

Virtually all antidepressant agents increase the birth of granule neurons in the adult dentate gyrus in rodents, providing a key basis for the neurogenesis hypothesis of antidepressant action. The novel antidepressant ketamine, however, shows antidepressant activity in humans within hours, far too rapid for a mechanism involving neuronal birth. Ketamine could potentially act more rapidly by enhancing maturation of new neurons born weeks earlier. To test this possibility, we assessed the effects of S-ketamine (S-(+)-ketamine hydrochloride) injection on maturation, as well as birth and survival, of new dentate gyrus granule neurons in rats, using the immediate-early gene zif268, proliferating cell nuclear antigen, and BrdU, respectively. We show that S-ketamine has rapid effects on new neurons, increasing the proportion of functionally mature young granule neurons within 2 h. A single injection of S-ketamine also increased cell proliferation and functional maturation, and decreased depressive-like behavior, for at least 4 weeks in rats treated with long-term corticosterone administration (a depression model) and controls. However, the behavioral effects of S-ketamine on neophagia were unaffected by elimination of adult neurogenesis. Together, these results indicate that ketamine has surprisingly rapid and long-lasting effects on the recruitment of young neurons into hippocampal networks, but that ketamine has antidepressant-like effects that are independent of adult neurogenesis.

Adult neurogenesis and mental illness.

Several lines of evidence suggest that adult neurogenesis, the production of new neurons in adulthood, may play a role in psychiatric disorders, including depression, anxiety, and schizophrenia. Medications and other treatments for mental disorders often promote the proliferation of new neurons; the time course for maturation and integration of new neurons in circuitry parallels the delayed efficacy of psychiatric therapies; adverse and beneficial experiences similarly affect development of mental illness and neurogenesis; and ablation of new neurons in adulthood alters the behavioral impact of drugs in animal models. At present, the links between adult neurogenesis and depression seem stronger than those suggesting a relationship between new neurons and anxiety or schizophrenia. Yet, even in the case of depression there is currently no direct evidence for a causative role. This article reviews the data relating adult neurogenesis to mental illness and discusses where research needs to head in the future.

Gap junctions in the ventral hippocampal-medial prefrontal pathway are involved in anxiety regulation.

Anxiety disorders are highly prevalent but little is known about their underlying mechanisms. Gap junctions exist in brain regions important for anxiety regulation, such as the ventral hippocampus (vHIP) and mPFC, but their functions in these areas have not been investigated. Using pharmacological blockade of neuronal gap junctions combined with electrophysiological recordings, we found that gap junctions play a role in theta rhythm in the vHIP and mPFC of adult mice. Bilateral infusion of neuronal gap junction blockers into the vHIP decreased anxiety-like behavior on the elevated plus maze and open field. Similar anxiolytic effects were observed with unilateral infusion of these drugs into the vHIP combined with contralateral infusion into the mPFC. No change in anxious behavior was observed with gap junction blockade in the unilateral vHIP alone or in the bilateral dorsal HIP. Since physical exercise is known to reduce anxiety, we examined the effects of long-term running on the expression of the neuronal gap junction protein connexin-36 among inhibitory interneurons and found a reduction in the vHIP. Despite this change, we observed no alteration in theta frequency or power in long-term runners. Collectively, these findings suggest that neuronal gap junctions in the vHIP-mPFC pathway are important for theta rhythm and anxiety regulation under sedentary conditions but that additional mechanisms are likely involved in running-induced reduction in anxiety.

Physical exercise prevents stress-induced activation of granule neurons and enhances local inhibitory mechanisms in the dentate gyrus.

Physical exercise is known to reduce anxiety. The ventral hippocampus has been linked to anxiety regulation but the effects of running on this subregion of the hippocampus have been incompletely explored. Here, we investigated the effects of cold water stress on the hippocampus of sedentary and runner mice and found that while stress increases expression of the protein products of the immediate early genes c-fos and arc in new and mature granule neurons in sedentary mice, it has no such effect in runners. We further showed that running enhances local inhibitory mechanisms in the hippocampus, including increases in stress-induced activation of hippocampal interneurons, expression of vesicular GABA transporter (vGAT), and extracellular GABA release during cold water swim stress. Finally, blocking GABAA receptors in the ventral hippocampus, but not the dorsal hippocampus, with the antagonist bicuculline, reverses the anxiolytic effect of running. Together, these results suggest that running improves anxiety regulation by engaging local inhibitory mechanisms in the ventral hippocampus.

Differential effects of stress and glucocorticoids on adult neurogenesis.

Stress is known to inhibit neuronal growth in the hippocampus. In addition to reducing the size and complexity of the dendritic tree, stress and elevated glucocorticoid levels are known to inhibit adult neurogenesis. Despite the negative effects of stress hormones on progenitor cell proliferation in the hippocampus, some experiences which produce robust increases in glucocorticoid levels actually promote neuronal growth. These experiences, including running, mating, enriched environment living, and intracranial self-stimulation, all share in common a strong hedonic component. Taken together, the findings suggest that rewarding experiences buffer progenitor cells in the dentate gyrus from the negative effects of elevated stress hormones. This chapter considers the evidence that stress and glucocorticoids inhibit neuronal growth along with the paradoxical findings of enhanced neuronal growth under rewarding conditions with a view toward understanding the underlying biological mechanisms.

Stress, stress hormones, and adult neurogenesis.

The dentate gyrus of the hippocampus continues to produce new neurons throughout adulthood. Adult neurogenesis has been linked to hippocampal function, including learning and memory, anxiety regulation and feedback of the stress response. It is thus not surprising that stress, which affects hippocampal function, also alters the production and survival of new neurons. Glucocorticoids, along with other neurochemicals, have been implicated in stress-induced impairment of adult neurogenesis. Paradoxically, increases in corticosterone levels are sometimes associated with enhanced adult neurogenesis in the dentate gyrus. In these circumstances, the factors that buffer against the suppressive influence of elevated glucocorticoids remain unknown; their discovery may provide clues to reversing pathological processes arising from chronic exposure to aversive stress.