Cholecystokinin (CCK) is the most abundant neuropeptide that broadly regulates the physiological status of animals. Here, we present a two-color laser theta burst stimulation (L-TBS) protocol for simultaneous activation of Schaffer collateral and perforant pathway in the hippocampus of CCK Cre mice. We describe steps for heterosynaptic long-term potentiation induction by L-TBS. This technique allows for the examination of the neurotransmitter roles in synaptic modulation and facilitates the exploration of pathological mechanisms in genetic models of brain disorders in mice. For complete details on the use and execution of this protocol, please refer to Su et al.1.

To investigate the influence of forepaw sensorimotor deprivation on memory and synaptic plasticity, Sprague-Dawley rats were divided into two groups: a sham-operated group and a group deprived of forepaw sensorimotor function by microsurgical operation at postnatal day 13 (PN13). Behavioral and electrophysiological studies were performed at PN25, PN35, PN45, and PN60. Open field test was used to assess the spontaneous locomotor activity. Morris water maze was used to evaluate spatial reference learning and memory. The long-term potentiation (LTP) in the medial perforant path--dentate gyrus (MPP-DG) pathway was examined with hippocampal slices. We found that forepaw sensorimotor deprivation did not affect spontaneous activity of the rats. However, spatial reference learning and memory were significantly impaired in their early life (PN25, PN35, and PN45). In accordance with the behavior results, LTP in MPP-DG pathway was significantly suppressed in their early life. These data demonstrated that forepaw sensorimotor deprivation led to the impairments on spatial memory via inducing pronounced deficits in the MPP-DG pathway to exhibit LTP, one of the major cellular mechanisms underlying learning and memory.

Hippocampal CA1 pyramidal neurons receive two excitatory glutamatergic synaptic inputs: their most distal dendritic regions in the stratum lacunosum-moleculare (SLM) are innervated by the perforant path (PP), originating from layer III of the entorhinal cortex, while their more proximal regions of the apical dendrites in the stratum radiatum (SR) are innervated by the Schaffer-collaterals (SC), originating from hippocampal CA3 neurons. Endocannabinoids (eCBs) are naturally occurring mediators capable of modulating both GABAergic and glutamatergic synaptic transmission and plasticity via the CB1 receptor. Previous work on eCB modulation of excitatory synapses in the CA1 region largely focuses on the SC pathway. However, little information is available on whether and how eCBs modulate glutamatergic synaptic transmission and plasticity at PP synapses.

In recent years, piperine has attracted much attention due to its various biological effects as a neuroprotective agent. Therefore, clarification of the possible side effects of piperine is important to identify its potential pharmacological action. Thus, the effects of piperine on the long-term plasticity of perforant pathway to dentate gyrus synapses were studied in hippocampus of an animal model of Alzheimer's disease (AD). Adult male rats were injected with intracerebroventricular (ICV) streptozotocin (STZ) bilaterally, on days 1 and 3 (3 mg/kg). The STZ-injected rats were treated with different doses of piperine for 4 weeks before being used in behavioral, electrophysiological and histopathological experiments. The passive-avoidance test was conducted on all animals in order to determine the cognitive performance. Rats were placed in a stereotaxic frame to implant a recording electrode in the hippocampal dentate gyrus and a stimulating electrode in the perforant path. Additionally, we assessed the density of survived neurons stained by cresyl violet. In this study, chronic administration of piperine low dose improved the ICV-STZ induced learning and long-term potentiation (LTP) impairments with no significant effect on baseline synaptic activity. In contrast, remarkable learning and long-term plasticity impairments were observed in rats treated by high dose of piperine in comparison to the other groups. Interestingly, this impaired hippocampal LTP was accompanied by an obvious alteration in baseline activity and significantly decreased neuronal numbers within the hippocampus. Therefore, our data provides a new understanding of the piperine supplementation effects on hippocampal electrophysiological profile although the consequences may be either beneficial or detrimental.

Hypertension is the most common chronic disease accompanied by cognitive decline and anxiety-like behavior. Angiotensin II (Ang II) induces hypertension by activating angiotensin II receptor subtype 1 (AT1R). The purpose of the study was to examine the potential underlying mechanism of alterations in cognition and anxiety-like behavior induced by Ang II. Adult C57 mice were intraperitoneal injected with either 1 mg/kg/d Ang II or saline individually for 14 consecutive days. Ang II resulted in cognitive decline and anxious like behavior in C57 mice. Moreover, Ang II disturbed bidirectional synaptic plasticity and neural oscillation coupling between high theta and gamma on PP (perforant pathway)-DG (dentate gyrus) pathway. In addition, Ang II decreased the expression of N-methyl-d-aspartate receptor (NR) 2A and NR 2B and increased the expression of GABAAR α1. The data suggest that Ang II disturb neural oscillations via altering excitatory and inhibitory (E/I) balance and eventually damage cognition and anxiety-like behavior in mice.

RBFOX3 mutations are linked to epilepsy and cognitive impairments, but the underlying pathophysiology of these disorders is poorly understood. Here we report replication of human symptoms in a mouse model with disrupted Rbfox3. Rbfox3 knockout mice displayed increased seizure susceptibility and decreased anxiety-related behaviors. Focusing on hippocampal phenotypes, we found Rbfox3 knockout mice showed increased expression of plasticity genes Egr4 and Arc, and the synaptic transmission and plasticity were defective in the mutant perforant pathway. The mutant dentate granules cells exhibited an increased frequency, but normal amplitude, of excitatory synaptic events, and this change was associated with an increase in the neurotransmitter release probability and dendritic spine density. Together, our results demonstrate anatomical and functional abnormality in Rbfox3 knockout mice, and may provide mechanistic insights for RBFOX3-related human brain disorders.

Mutations in the putative glutamatergic synapse scaffolding protein SAP97 are associated with the development of schizophrenia in humans. However, the role of SAP97 in synaptic regulation is unclear. Here we show that SAP97 is expressed in the dendrites of granule neurons in the dentate gyrus but not in the dendrites of other hippocampal neurons. Schizophrenia-related perturbations of SAP97 did not affect CA1 pyramidal neuron synapse function. Conversely, these perturbations produce dramatic augmentation of glutamatergic neurotransmission in granule neurons that can be attributed to a release of perisynaptic GluA1-containing AMPA receptors into the postsynaptic densities of perforant pathway synapses. Furthermore, inhibiting SAP97 function in the dentate gyrus was sufficient to impair contextual episodic memory. Together, our results identify a cell-type-specific synaptic regulatory mechanism in the dentate gyrus that, when disrupted, impairs contextual information processing in rats.

A prospective, standardised neuroimaging protocol was implemented to characterise mesial temporal lobe pathology in amyotrophic lateral sclerosis, Alzheimer's disease and healthy controls focusing on the evaluation of interconnected white and grey matter structures. "Hippocampal pathology in Amyotrophic Lateral Sclerosis: selective vulnerability of subfields and their associated projections" [1]. High-resolution diffusion tensor and structural imaging data were acquired on a 3 T MRI platform using standardised sequence parameters. The integrity of the fornix and the perforant pathway was assessed by tractography, to provide fractional anisotropy, axial diffusivity and radial diffusivity measures. Quantitative structural imaging was used to estimate the total intracranial volume, total hippocampal volumes and hippocampal subfield volumes for each participant. Raw white- and grey-matter measures, demographic and clinical data are available online at 'Mendeley Data'. Amyotrophic lateral sclerosis and Alzheimer's disease exhibit divergent hippocampal profiles.

While early 1990s reports showed the phosphorylation pattern of fetal tau protein to be similar to that of tau in paired helical filaments (PHF) in Alzheimer's disease (AD), neither the molecular mechanisms of the transient developmental hyperphosphorylation of tau nor reactivation of the fetal plasticity due to re-expression of fetal protein kinases in the aging and AD human brain have been sufficiently investigated. Here, we summarize the current knowledge on fetal tau, adding new data on the specific patterns of tau protein and mRNA expression in the developing human brain as well as on change in tau phosphorylation in the perforant pathway after entorhinal cortex lesion in mice. As fetal tau isoform does not form PHF even in a highly phosphorylated state, understanding its expression and post-translational modifications represents an important avenue for future research towards the development of AD treatment and prevention.

The presubiculum (PRS) is an integral component of the perforant pathway that has recently been recognised as a relatively unscathed region in clinical Alzheimer's disease (AD), despite neighbouring components of the perforant pathway, CA1 and the entorhinal cortex, responsible for formation of episodic memory and storage, showing severe hallmarks of AD including, amyloid-beta (Aβ) plaques, tau tangles and marked gliosis. However, the question remains whether this anatomical resilience translates into functional resilience of the PRS neurons. Using neuroanatomy combined with whole-cell electrophysiological recordings, we investigated whether the unique spatial profile of the PRS was replicable in two knock-in mouse models of AD, APPNL-F/NL-F, and APPNL-F/MAPTHTAU and whether the intrinsic properties and morphological integrity of the PRS principal neurons was maintained compared to the lateral entorhinal cortex (LEC) and hippocampal CA1 principal cells. Our data revealed an age-dependent Aβ and tau pathology with neuroinflammation in the LEC and CA1, but a presence of fleece-like Aβ deposits with an absence of tau tangles and cellular markers of gliosis in the PRS of the mouse models at 11-16 and 18-22 months. These observations were consistent in human post-mortem AD tissue. This spatial profile also correlated with functional resilience of strong burst firing PRS pyramidal cells that showed unaltered sub- and suprathreshold intrinsic biophysical membrane properties and gross morphology in the AD models that were similar to the properties of pyramidal cells recorded in age-matched wild-type mice (11-14 months). This was in contrast to the LEC and CA1 principal cells which showed altered subthreshold intrinsic properties such as a higher input resistance, longer membrane time constants and hyperexcitability in response to suprathreshold stimulation that correlated with atrophied dendrites in both AD models. In conclusion, our data show for the first time that the unique anatomical profile of the PRS constitutes a diffuse AD pathology that is correlated with the preservation of principal pyramidal cell intrinsic biophysical and morphological properties despite alteration of LEC and CA1 pyramidal cells in two distinct genetic models of AD. Understanding the underlying mechanisms of this resilience could be beneficial in preventing the spread of disease pathology before cognitive deficits are precipitated in AD.

Frequency-dependent hippocampal activation during electrical perforant pathway stimulation was analyzed simultaneously by electrophysiological recordings in dentate gyrus and functional magnetic resonance imaging (fMRI). Pulse trains at low-frequency stimulation (2.5 Hz) did not influence electrophysiological responses within stimulation trains in the dentate gyrus and triggered no detectable BOLD responses. Increased stimulation frequencies (5.0-20 Hz) generated a roughly linear enhancement of the BOLD response. The BOLD signal within the dentate gyrus correlated more closely with stimulus pattern than with generated action potentials of the granular cells. However, the BOLD signal was strongly influenced by additional local signal processing activated by repetitive stimulus trains. fMRI visualized a frequency-specific spatial activation pattern of the hippocampus; spatially restricted activation in the dentate gyrus during 5-Hz stimulation, activation of the entire hippocampus and subiculum at 10 Hz and activation of the contralateral hippocampus during 20-Hz stimulation.

This study focused on the effects of zinc oxide nanoparticles (nano-ZnO) on spatial learning and memory and synaptic plasticity in the hippocampus of young rats, and tried to interpret the underlying mechanism. Rats were randomly divided into four groups. Nano-ZnO and phosphate-buffered saline were administered in 4-week-old rats for 8 weeks. Subsequently, performance in Morris water maze (MWM) was determined, and then long-term potentiation (LTP) and depotentiation were measured in the perforant pathway to dentate gyrus (DG) in anesthetized rats. The data showed that, (1) in MWM, the escape latency was prolonged in the nano-ZnO group and, (2) LTP was significantly enhanced in the nano-ZnO group, while depotentiation was barely influenced in the DG region of the nano-ZnO group. This bidirectional effect on long-term synaptic plasticity broke the balance between stability and flexibility of cognition. The spatial learning and memory ability was attenuated by the alteration of synaptic plasticity in nano-ZnO-treated rats.

We used a 24 h perforant path stimulation model of status epilepticus to study the role of non-NMDA receptors in the loss of hilar interneurons and paired pulse inhibition associated with the model. In one experiment, NBQX administered i.v. at 1.0 mg/kg/h significantly reduced the loss of hematoxylin and eosin-stained hilar neurons from 360.2 to 125.3 but failed to protect against the loss of paired pulse inhibition. In a second experiment, i.v. NBQX at 1.5 mg/kg/h significantly protected against loss of SS- and NPY-positive hilar interneurons but also failed to protect against loss of paired pulse inhibition. These results demonstrate that the neuronal loss associated with sustained stimulation of this excitatory pathway is mediated in part through non-NMDA receptors. The lack of protection against loss of paired pulse inhibition suggests that SS- and NPY-immunoreactive interneurons may not be responsible for frequency-dependent paired-pulse inhibition of dentate granule cells.

Memory impairment is a common after an ischemic stroke. While delayed neuronal death in the CA1 region is usually linked to cerebral ischemia-induced memory impairment, the role of early immature neuronal death within the DG region in the memory state of an ischemic stroke model has rarely been studied. Here, we show a partial role of immature neuronal death in memory impairment in a global ischemia model. We found early immature neuronal death, which was determined by DCX and NeuN-double-staining. Injection of z-DEVD-fmk, a caspase-3 inhibitor, into the DG region rescued cells from immature neuronal death in the DG region without affecting delayed neuronal death in the CA1 region of an ischemic brain. Moreover, z-DEVD-fmk treatment partially rescued ischemia-induced spatial memory impairment. We also found that ischemia-induced LTP impairment in the perforant pathway was restored by z-DEVD-fmk treatment. These results suggest that early immature neuronal death is partially involved in ischemia-induced spatial memory impairment.

While hippocampal connectivity is essential to normal memory function, our knowledge of human hippocampal circuitry is largely inferred from animal studies. Using polarized light microscopy at 1.3 µm resolution, we have directly visualized the 3D course of key medial temporal pathways in 3 ex vivo human hemispheres and 2 ex vivo vervet monkey hemispheres. The multiple components of the perforant path system were clearly identified: Superficial sheets of fibers emanating from the entorhinal cortex project to the presubiculum and parasubiculum, intermixed transverse and longitudinal angular bundle fibers perforate the subiculum and then project to the cornu ammonis (CA) fields and dentate molecular layer, and a significant alvear component runs from the angular bundle to the CA fields. From the hilus, mossy fibers localize to regions of high kainate receptor density, and the endfolial pathway, mostly investigated in humans, merges with the Schaffer collaterals. This work defines human hippocampal pathways underlying mnemonic function at an unprecedented resolution.

The hippocampus receives glutamatergic and GABAergic inputs from subcortical regions. Despite the important roles of these subcortical inputs in the regulation of hippocampal circuit, it has not been explored whether associative activation of the subcorticohippocampal pathway induces Hebbian plasticity of subcortical inputs. Here, we demonstrate that the hypothalamic supramammillary nucleus (SuM) to the dentate granule cell (GC) synapses, which co-release glutamate and GABA, undergo associative long-term potentiation (LTP) of glutamatergic, but not GABAergic, co-transmission. This LTP is induced by pairing of SuM inputs with GC spikes. We found that this Hebbian LTP is input-specific, requires NMDA receptors and CaMKII activation, and is expressed postsynaptically. By the net increase in excitatory drive of SuM inputs following LTP induction, associative inputs of SuM and the perforant path effectively discharge GCs. Our results highlight the important role of associative plasticity at SuM-GC synapses in the regulation of dentate gyrus activity and for the encoding of SuM-related information.

The recent discovery of a large latent population of precursor cells in the dentate gyrus of adult mice led us to investigate whether activation of this population is regulated by synaptic activity, thereby explaining the observation that environmental signals can affect neurogenesis. Using a variety of stimulation protocols, we found that only a long-term potentiation (LTP)-inducing protocol activated the latent precursor pool, leading to increased neurogenesis in the dentate gyrus. LTP induced by high-frequency stimulation (HFS) of the perforant pathway in vivo produced a two-fold increase in the number of neurospheres cultured from the stimulated hippocampus, compared with the unstimulated hippocampus. No increase in neurosphere number or neurogenesis was observed when the HFS failed to induce LTP. These results show that LTP can activate latent neural precursor cells in the adult mouse dentate gyrus, thereby providing a direct mechanism for regulating activity-driven neurogenesis. In the future, it may be possible to utilize such learning- or stimulation-induced neurogenesis to overcome disorders characterized by neuronal loss.

In Alzheimer's disease (AD), a decline in explicit memory is one of the earliest signs of disease and is associated with hippocampal dysfunction. Amyloid protein exerts a disruptive impact on neuronal function, but the specific effects on hippocampal network activity are not well known. In this study, fast voltage-sensitive dye imaging and extracellular and whole-cell electrophysiology were used on entorhinal cortical-hippocampal slice preparations to characterize hippocampal network activity in 12-16 month old female APPswe/PSEN1DeltaE9 (APdE9 mice) mice. Aged APdE9 mice exhibited profound disruptions in dentate gyrus circuit activation. High frequency stimulation of the perforant pathway in the dentate gyrus (DG) area of APdE9 mouse tissue evoked abnormally large field potential responses corresponding to the wider neural activation maps. Whole-cell patch clamp recordings of the identified inhibitory interneurons in the molecular layer of DG revealed that they fail to reliably fire action potentials. Taken together, abnormal DG excitability and an inhibitory neuron failure to generate action potentials are suggested to be important contributors to the underlying cellular mechanisms of early-stage Alzheimer's disease pathophysiology.

The brain circuits and synaptic processes that underlie alcohol addiction are currently the subject of intensive research. Here we focus on hippocampal circuitry and show that chemogenetic inhibition of dentate gyrus (DG) during presentation of alcohol-associated cues has long-lasting effects on mice behavior. DG inhibition enhances alcohol seeking and drinking, suggesting that DG regulates addiction-related behaviors. To test this hypothesis, we perform whole-cell patch-clamp recordings from the granule cells of DG and look for electrophysiological correlates of alcohol addiction. We observe that presentation of alcohol-associated cue light that induces relapse to alcohol-seeking results in generation of silent synapses, that lack functional AMPA receptors. Furthermore, using human criteria of addiction, we differentiate mice controlling their alcohol consumption from those that undergo transition to addiction to discover that the levels of silent synapses induced by alcohol cues are specifically increased in the addicted mice. As the total level of dendritic spines that harbor synapses is constant at this time point, our data indicate that synapses of perforant path to DG are weakened during cue relapse. Finally we demonstrate that, acamprosate, a drug that limits alcohol drinking and seeking in addicts, prevents generation of silent synapses in DG upon presentation of alcohol-associated cues. Altogether, our data suggest that weakening of DG synapses upon cue relapse contributes to persistent alcohol addiction-related behaviors.

Increasing evidence suggests that long-term consumption of high-caloric diets increases the risk of developing cognitive dysfunctions. In the present study, we assessed the catecholaminergic activity in the hippocampus as a modulatory mechanism that is altered in rats exposed to six months of a high-sucrose diet (HSD). Male Wistar rats fed with this diet developed a metabolic disorder and showed impaired spatial memory in both water maze and object location memory (OLM) tasks. Intrahippocampal free-movement microdialysis showed a diminished dopaminergic and noradrenergic response to object exploration during OLM acquisition compared to rats fed with normal diet. In addition, electrophysiological results revealed an impaired long-term potentiation (LTP) of the perforant to dentate gyrus pathway in rats exposed to a HSD. Local administration of nomifensine, a catecholaminergic reuptake inhibitor, prior to OLM acquisition or LTP induction, improved long-term memory and electrophysiological responses, respectively. These results suggest that chronic exposure to HSD induces a hippocampal deterioration which impacts on cognitive and neural plasticity events negatively; these impairments can be ameliorated by increasing or restituting the affected catecholaminergic activity.