The septotemporal axis of the hippocampus separates it into domains with unique molecular, cellular, downstream connectivity, and behavioral profiles, and yet very little is known about the ontogenesis of these highly specialized subcircuits. Here, we use viral tracing, optogenetic-assisted patch clamping, chemogenetics, and behavior in mice to examine changes in domain-defined hippocampus efferent projections from postnatal day (P)10 to P60. We find distinct anatomical and synaptic developmental signatures in ventral and intermediate CA1 downstream connectivity, with unique contributions to the prelimbic and infralimbic subregions of the medial prefrontal cortex (mPFC). Juvenile inhibition of the ventral and intermediate CA1-mPFC pathways leads to opposing modulation of adult cognitive flexibility, establishing a sex- and pathway-specific sensitive period preceding the stabilization of CA1-mPFC synaptic transmission. Our data elucidate domain- and target-defined postnatal maturation of hippocampus efferents, indicating juvenility as a CA1-mPFC sensitive period with crucial implications for early life influences on adult cognition.

Delay discounting (DD) is a phenomenon where individuals devalue a reward associated with a temporal delay, with the rate of devaluation being representative of impulsive-like behavior. Here, we first sought to develop and validate a mouse DD task to study brain circuits involved in DD decision-making within short developmental time windows, given widespread evidence of developmental regulation of impulse control and risk-taking. We optimized a T-maze DD task for mice that enables training and DD trials within 2 weeks. Mice learned to choose between a large and a small reward located at opposite arms of a T-maze. Once training criteria were met, mice underwent DD whereby the large reward choice was associated with a temporal delay. Task validation showed that adolescent C57BL/6J mice display an increased preference for the small reward upon a temporal delay, confirming increased impulsivity compared with adults. We next used this DD task to explore the neural basis of decision-making. We used tyrosine hydroxylase transgenic mice (TH-Cre) to target TH-positive neurons in the nucleus accumbens (NAc) and ventral tegmental area (VTA) with Cre-dependent excitatory or inhibitory designer receptors exclusively activated by designer drugs (DREADDs). Inhibition of transduced neurons in the NAc decreased preference for the small but immediate reward during DD. Inhibition of TH-positive neurons in the ventral tegmental area (VTA) did not affect impulsive choice in this DD task. These results uncover a novel role for NAc TH-positive neurons in DD behavior and expand the repertoire of behavioral tasks available for studying decision-making across the lifespan.

The ventral hippocampus (vHPC) and its projections are key regulators of affective behavior and mood and anxiety disorders. While approximately half of all lifetime cases of mood and anxiety disorders start by adolescence, very little is known about how vHPC efferent circuits form and regulate behavior during early life. Here we used a combination of optogenetics, electrophysiology and behavior to map the synaptic and anatomical maturation of vHIP efferent circuits and their implications for the ontogeny of behavior.

The ability to generate memories that persist throughout a lifetime (that is, memory persistence) emerges in early development across species. Although it has been shown that persistent fear memories emerge between late infancy and adolescence in mice, it is unclear exactly when this transition takes place, and whether two major fear conditioning tasks, contextual and auditory fear, share the same time line of developmental onset. Here, we compared the ontogeny of remote contextual and auditory fear in C57BL/6J mice across early life. Mice at postnatal day (P)15, 21, 25, 28, and 30 underwent either contextual or auditory fear training and were tested for fear retrieval 1 or 30 d later. We found that mice displayed 30-d memory for context– and tone–fear starting at P25. We did not find sex differences in the ontogeny of either type of fear memory. Furthermore, 30-d contextual fear retrieval led to an increase in the number of c-Fos positive cells in the prelimbic region of the prefrontal cortex only at an age in which the contextual fear memory was successfully retrieved. These data delineate a precise time line for the emergence of persistent contextual and auditory fear memories in mice and suggest that the prelimbic cortex is only recruited for remote memory recall upon the onset of memory persistence.

Adeno-associated viruses (AAVs) are a commonly used tool in neuroscience to efficiently label, trace, and/or manipulate neuronal populations. Highly specific targeting can be achieved through recombinase-dependent AAVs in combination with transgenic rodent lines that express Cre-recombinase in specific cell types. Visualization of viral expression is typically achieved through fluorescent reporter proteins (e.g., GFP or mCherry) packaged within the AAV genome. Although nonamplified fluorescence is usually sufficient to observe viral expression, immunohistochemical amplification of the fluorescent reporter is routinely used to improve viral visualization. In the present study, Cre-dependent AAVs were injected into the neocortex of wild-type C57BL/6J mice. While we observed weak but consistent nonamplified off-target double inverted open reading frame (DIO) expression in C57BL/6J mice, antibody amplification of the GFP or mCherry reporter revealed notable Cre-independent viral expression. Off-target expression of DIO constructs in wild-type C57BL/6J mice occurred independent of vendor, AAV serotype, or promoter. We also evaluated whether Cre-independent expression had functional effects via designer receptors exclusively activated by designer drugs (DREADDs). The DREADD agonist C21 (compound 21) had no effect on contextual fear conditioning or c-Fos expression in DIO-hM3Dq-mCherry(+) cells of C57BL/6J mice. Together, our results indicate that DIO constructs have off-target expression in wild-type subjects. Our findings are particularly important for the design of experiments featuring sensitive systems and/or quantitative measurements that could be negatively impacted by off-target expression.

Spontaneous recognition memory tasks explore the what, where and when components of recognition memory. These tasks are widely used in rodents to assess cognitive function across the lifespan. While several neurodevelopmental and mental disorders present symptom onset in early life, very little is known about how memories are expressed in early life, and as a consequence how they may be affected in pathological conditions. In this review, we conduct an analysis of the studies examining the expression of spontaneous recognition memory in young rodents. We compiled studies using four different tasks: novel object recognition, object location, temporal order recognition and object place. First, we identify major sources of variability between early life spontaneous recognition studies and classify them for later comparison. Second, we use these classifications to explore the current knowledge on the ontogeny of each of these four spontaneous recognition memory tasks. We conclude by discussing the possible implications of the relative time of onset for each of these tasks and their respective neural correlates. In compiling this research, we hope to advance on establishing a developmental timeline for the emergence of distinct components of recognition memory, while also identifying key areas of focus for future research. Establishing the ontogenetic profile of rodent spontaneous recognition memory tasks will create a necessary blueprint for cognitive assessment in animal models of neurodevelopmental and mental disorders, a first step towards improved and earlier diagnosis as well as novel intervention strategies.

Spontaneous recognition memory tasks build on an animal’s natural preference for novelty to assess the what, where and when components of episodic memory. Their simplicity, ethological relevance and cross-species adaptability make them extremely useful to study the physiology and pathology of memory. Recognition memory deficits are common in rodent models of neurodevelopmental disorders, and yet very little is known about the expression of spontaneous recognition memory in young rodents. This is exacerbated by the paucity of data on the developmental onset of recognition memory in mice, a major animal model of disease. To address this, we characterized the ontogeny of three types of spontaneous recognition memory in mice: object location, novel object recognition and temporal order recognition. We found that object location is the first to emerge, at postnatal day (P)21. This was followed by novel object recognition (24 h delay), at P25. Temporal order recognition was the last to emerge, at P28. Elucidating the developmental expression of recognition memory in mice is critical to improving our understanding of the ontogeny of episodic memory, and establishes a necessary blueprint to apply these tasks to probe cognitive deficits at clinically relevant time points in animal models of developmental disorders.

Early life experiences critically define cognitive and mental health function throughout life. Childhood and adolescence are the predominant age of onset for the majority of mental disorders, periods in which key brain areas involved in emotional processing, such as medial prefrontal cortex (mPFC) and amygdala, are maturing. Anatomical and morphological changes occur in both these areas during early life; nevertheless, how these changes affect circuit function and its consequences to the onset of mental illness is currently unknown.

A brain network comprising the medial prefrontal cortex (mPFC) and amygdala plays important roles in developmentally regulated cognitive and emotional processes. However, very little is known about the maturation of mPFC-amygdala circuitry. We conducted anatomical tracing of mPFC projections and optogenetic interrogation of their synaptic connections with neurons in the basolateral amygdala (BLA) at neonatal to adult developmental stages in mice. Results indicate that mPFC-BLA projections exhibit delayed emergence relative to other mPFC pathways and establish synaptic transmission with BLA excitatory and inhibitory neurons in late infancy, events that coincide with a massive increase in overall synaptic drive. During subsequent adolescence, mPFC-BLA circuits are further modified by excitatory synaptic strengthening as well as a transient surge in feedforward inhibition. The latter was correlated with increased spontaneous inhibitory currents in excitatory neurons, suggesting that mPFC-BLA circuit maturation culminates in a period of exuberant GABAergic transmission. These findings establish a time course for the onset and refinement of mPFC-BLA transmission and point to potential sensitive periods in the development of this critical network.SIGNIFICANCE STATEMENT Human mPFC-amygdala functional connectivity is developmentally regulated and figures prominently in numerous psychiatric disorders with a high incidence of adolescent onset. However, it remains unclear when synaptic connections between these structures emerge or how their properties change with age. Our work establishes developmental windows and cellular substrates for synapse maturation in this pathway involving both excitatory and inhibitory circuits. The engagement of these substrates by early life experience may support the ontogeny of fundamental behaviors but could also lead to inappropriate circuit refinement and psychopathology in adverse situations.

The ability to form associations between aversive threats and their predictors is fundamental to survival. However, fear and anxiety in excess are detrimental and are a hallmark of psychiatric diseases such as post-traumatic stress disorder (PTSD). PTSD symptomatology includes persistent and intrusive thoughts of an experienced trauma, suggesting an inability to downregulate fear when a corresponding threat has subsided. Convergent evidence from human and rodent studies supports a role for the medial prefrontal cortex (mPFC)-amygdala network in both PTSD and the regulation of fear memory expression. In particular, current models stipulate that the prelimbic (PL) and infralimbic (IL) subdivisions of the rodent mPFC bidirectionally regulate fear expression via differential recruitment of amygdala neuronal subpopulations. However, an array of recent studies that employ new technical approaches has fundamentally challenged this interpretation. Here we explore how a new emphasis on the contribution of inhibitory neuronal populations, subcortical structures and the passage of time is reshaping our understanding of mPFC-amygdala circuits and their control over fear.

Proliferation of neural progenitor cells in the subventricular zone leads to the continuous generation of new olfactory granule cells (OGCs) throughout life. These cells synaptically integrate into olfactory bulb circuits after ∼2 weeks and transiently exhibit heightened plasticity and responses to novel odors. Although these observations suggest that adult-generated OGCs play important roles in olfactory-related memories, global suppression of olfactory neurogenesis does not typically prevent the formation of odor–reward memories, perhaps because residual OGCs can compensate. Here, we used a transgenic strategy to selectively ablate large numbers of adult-generated OGCs either before or after learning in mice. Consistent with previous studies, pretraining ablation of adult-generated OGCs did not prevent the formation of an odor–reward memory, presumably because existing OGCs can support memory formation in their absence. However, ablation of a similar cohort of adult-generated OGCs after training impaired subsequent memory expression, indicating that if these cells are available at the time of training, they play an essential role in subsequent expression of odor–reward memories. Memory impairment was associated with the loss of adult-generated OGCs that were >10 d in age and did not depend on the developmental stage in which they were generated, suggesting that, once sufficiently mature, OGCs generated during juvenility and adulthood play similar roles in the expression of odor–reward memories. Finally, ablation of adult-generated OGCs 1 month after training did not produce amnesia, indicating that adult-generated OGCs play a time-limited role in the expression of odor–reward memories.

Conditioned fear requires neural activity in the basolateral amygdala (BLA) and medial prefrontal cortex (mPFC), structures that are densely interconnected at the synaptic level. Previous work has suggested that anatomical subdivisions of mPFC make distinct contributions to fear expression and inhibition, and that the functional output of this processing is relayed to the BLA complex. However, it remains unknown whether synaptic plasticity in mPFC-BLA networks contributes to fear memory encoding. Here we use optogenetics andex vivoelectrophysiology to reveal the impact of fear conditioning on BLA excitatory and feedforward inhibitory circuits formed by projections from infralimbic (IL) and prelimbic (PL) cortices. In naive mice, these pathways recruit equivalent excitation and feedforward inhibition in BLA principal neurons. However, fear learning leads to a selective decrease in inhibition:excitation balance in PL circuits that is attributable to a postsynaptic increase in AMPA receptor function. These data suggest a pathway-specific mechanism for fear memory encoding by adjustment of mPFC-BLA transmission. Upon reengagement of PL by conditioned cues, these modifications may serve to amplify emotional responses.

New granule cells are continuously integrated into hippocampal circuits throughout adulthood, and the fine-tuning of this process is likely important for efficient hippocampal function. During development, this integration process is critically regulated by the α-calcium/calmodulin-dependent protein kinase II (α-CaMKII), and here we ask whether this role is conserved in the adult brain. To do this, we developed a transgenic strategy to conditionally delete α-CaMKII from neural progenitor cells and their progeny in adult mice. First, we found that the selective deletion of α-CaMKII from newly generated dentate granule cells led to an increase in dendritic complexity. Second, α-CaMKII deletion led to a reduction in number of mature synapses and cell survival. Third, consistent with altered morphological and synaptic development, acquisition of one-trial contextual fear conditioning was impaired after deletion of α-CaMKII from newly generated dentate granule cells. Previous work in Xenopus identified α-CaMKII as playing a key role in the stabilization of dendritic and synaptic structure during development. The current study indicates that α-CaMKII plays a plays a similar, cell-autonomous role in the adult hippocampus and, in addition, reveals that the loss of α-CaMKII from adult-generated granule cells is associated with impaired hippocampus-dependent learning.

Pinpointing the precise age when young animals begin to form memories of aversive events is valuable for understanding the onset of anxiety and mood disorders and for detecting early cognitive impairment in models of childhood-onset disorders. Although these disorders are most commonly modeled in mice, we know little regarding the development of learning and memory in this species because most previous studies have been restricted to rats. Therefore, in the present study, we constructed an ontogenetic timeline of contextual fear memory ranging from infancy to adulthood in mice. We found that the ability of mice to form long-term context-shock associations emerged ∼13-14 d of age, which is several days earlier than previously reported for rats. Although the ability to form contextual fear memories remained stable from infancy into adulthood, infant mice had shorter-lasting memories than adolescent and adult mice. Furthermore, we found that mice subjected to fetal alcohol exposure showed a delay in the developmental emergence of contextual fear memory, illustrating the utility of this ontogenetic approach in detecting developmental delays in cognitive function stemming from maladaptive early life experience.

Newly generated dentate granule cells in the hippocampus at 4 weeks after their 'birth' are more plastic than existing neurons. The authors use a combined retroviral and optogenetic approach to show that silencing these 4-week-old cells, but not cells of other ages, impaired retrieval of hippocampal memory. Accumulating evidence suggests that global depletion of adult hippocampal neurogenesis influences its function and that the timing of the depletion affects the deficits. However, the behavioral roles of adult-born neurons during their establishment of projections to CA3 pyramidal neurons remain largely unknown. We used a combination of retroviral and optogenetic approaches to birth date and reversibly control a group of adult-born neurons in adult mice. Adult-born neurons formed functional synapses on CA3 pyramidal neurons as early as 2 weeks after birth, and this projection to the CA3 area became stable by 4 weeks in age. Newborn neurons at this age were more plastic than neurons at other stages. Notably, we found that reversibly silencing this cohort of ∼4-week-old cells after training, but not cells of other ages, substantially disrupted retrieval of hippocampal memory. Our results identify a restricted time window for adult-born neurons essential in hippocampal memory retrieval.

Galectins are a 15 member family of carbohydrate-binding proteins that have been implicated in cancer, immunity, inflammation and development. While galectins are expressed in the central nervous system, little is known about their function in the adult brain. Previously we have shown that galectin-1 (gal-1) is expressed in the adult hippocampus, and, in particular, in putative neural stem cells in the subgranular zone. To evaluate how gal-1 might contribute to hippocampal memory function here we studied galectin-1 null mutant (gal-1-/-) mice. Compared to their wildtype littermate controls, gal-1-/- mice exhibited impaired spatial learning in the water maze and contextual fear learning. Interestingly, tone fear conditioning was normal in gal-1-/- mice suggesting that loss of gal-1 might especially impact hippocampal learning and memory. Furthermore, gal-1-/- mice exhibited normal motor function, emotion and sensory processing in a battery of other behavioral tests, suggesting that non-mnemonic performance deficits are unlikely to account for the spatial and contextual learning deficits. Together, these data reveal a role for galectin-carbohydrate signalling in hippocampal memory function.

New neurons are continuously generated in the subgranular zone of the adult hippocampus and, once sufficiently mature, are thought to integrate into hippocampal memory circuits. However, whether they play an essential role in subsequent memory expression is not known. Previous studies have shown that suppression of adult neurogenesis often (but not always) impairs subsequent hippocampus-dependent learning (i.e., produces anterograde effects). A major challenge for these studies is that these new neurons represent only a small subpopulation of all dentate granule cells, and so there is large potential for either partial or complete compensation by granule cells generated earlier on during development. A potentially more powerful approach to investigate this question would be to ablate adult-generated neurons after they have already become part of a memory trace (i.e., retrograde effects). Here we developed a diphtheria toxin-based strategy in mice that allowed us to selectively ablate a population of predominantly mature, adult-generated neurons either before or after learning, without affecting ongoing neurogenesis. Removal of these neurons before learning did not prevent the formation of new contextual fear or water maze memories. In contrast, removal of an equivalent population after learning degraded existing contextual fear and water maze memories, without affecting nonhippocampal memory. Ablation of these adult-generated neurons even 1 month after learning produced equivalent memory degradation in the water maze. These retrograde effects suggest that adult-generated neurons form a critical and enduring component of hippocampal memory traces.

Processing olfactory information is critical for an animal's ability to locate food, detect danger, recognize conspecifics, find mates, and rear offspring. The olfactory bulb (OB) is the initial site of olfactory information processing in the brain. Within the OB, mitral cells receive input from