Differential effects of aging on hippocampal ultrastructure in male vs. female rats.

Age-related decline in physical and cognitive functions are facts of life that do not affect everyone to the same extent. We had reported earlier that such cognitive decline is both sex- and context-dependent. Moreover, age-associated ultrastructural changes were observed in the hippocampus of male rats. In this study, we sought to determine potential differences in ultrastructural changes between male and female rats at various stages of life. We performed quantitative electron microscopic evaluation of hippocampal CA1 region, an area intimately involved in cognitive behavior, in both male and female adolescent, adult and old Wistar rats. Specifically, we measured the number of docking synaptic vesicles in axo-dendritic synapses, the length of active zone as well as the total number of synaptic vesicles. Distinct age- and sex-dependent effects were observed in several parameters. Thus, adult female rats had the lowest synaptic active zone compared to both adolescent and old female rats. Moreover, the same parameter was significantly lower in adult and old female rats compared to their male counterparts. On the other hand, old male rats had significantly lower number of total synaptic vesicles compared to both adolescent and adult male rats as well as compared to their female counterparts. Taken together, it may be suggested that age- and sex-dependent ultrastructural changes in the hippocampus may underlie at least some of the differences in cognitive functions among these groups.

Increasing the interval between repeated anesthetic exposures reduces long-lasting synaptic changes in late post-natal mice.

While recent studies strongly suggest that a single, short anesthetic exposure does not affect neurodevelopment, the effects of multiple exposures remain unclear. Unfortunately, studying "multiple exposures" is challenging as it is an extremely heterogeneous descriptor comprising diverse factors. One potentially important, but unrecognized factor is the interval between anesthetic exposures. In order to evaluate the significance of interval, we exposed post-natal day 16, 17 mice to three sevoflurane exposures (2.5%, 1 hr) with short (2 hr) or long (24 hr) intervals. Changes in synaptic transmission, plasticity, protein expression, and behavior were assessed in male and female mice. We discovered that short-interval exposures induced a female-dependent decrease in miniature inhibitory post-synaptic current (mIPSC) frequency 5 days after the last exposure (control: 18.44 ± 2.86 Hz, sevoflurane:14.65 ± 4.54 Hz). Short-interval sevoflurane exposed mice also displayed long-term behavioral deficits at adult age (hypoactivity, anxiety). These behavioral changes were consistent with the sex-dependent changes in inhibitory transmission, as they were more robust in female mice. Although there was no change in learning and memory, short-interval sevoflurane exposures also impaired LTP in a non-sex-dependent manner (control: 171.10 ± 26.90%, sevoflurane: 149.80 ± 26.48 %). Most importantly, we were unable to find long-lasting consequences in mice that received long-interval sevoflurane exposures. Our study provides novel insights regarding the significance of the interval between multiple exposures, and also suggests that the neurotoxic effects of multiple anesthetic exposures may be reduced by simply increasing the interval between each exposure.

Activation of ventral CA1 hippocampal neurons projecting to the lateral septum during feeding.

A number of studies have reported the involvement of the ventral hippocampus (vHip) and the lateral septum (LS) in negative emotional responses. Besides these well-documented functions, they are also thought to control feeding behavior. In particular, optogenetic and pharmacogenetic interventions to LS-projecting vHip neurons have demonstrated that the vHip→LS neural circuit exerts an inhibition on feeding behavior. However, there have been no reports of vHip neuronal activity during feeding. Here, we focused on LS-projecting vCA1 neurons (vCA1→LS ) and monitored their activity during feeding behaviors in mice. vCA1→LS neurons were retrogradely labeled with adeno-associated virus carrying a ratiometric Ca2+ indicator and measured compound Ca2+ dynamics by fiber photometry. We first examined vCA1→LS activity in random food-exploring behavior and found that vCA1→LS activation seemed to coincide with food intake; however, our ability to visually confirm this during freely moving behaviors was not sufficiently reliable. We next examined vCA1→LS activity in a goal-directed, food-seeking lever-press task which temporally divided the mouse state into preparatory, effort, and consummatory phases. We observed vCA1→LS activation in the postprandial period during the consummatory phase. Such timing- and pathway-specific activation was not observed from pan-vCA1 neurons. In contrast, reward omission eliminated this activity, indicating that vCA1→LS activation is contingent on the food reward. Sated mice pressed the lever significantly fewer times but still ate food; however, vCA1→LS neurons were not activated, suggesting that vCA1→LS neurons did not respond to habitual behavior. Combined, these results suggest that gastrointestinal interoception rather than food-intake motions or external sensations are likely to coincide with vCA1→LS activity. Accordingly, we propose that vCA1→LS neurons discriminate between matched or unmatched predictive bodily states in which incoming food will satisfy an appetite. We also demonstrate that vCA1→LS neurons are activated in aversive/anxious situations in an elevated plus maze and tail suspension test. Future behavioral tests utilizing anxious conflict and food intake may reconcile the multiple functions of vCA1→LS neurons.

Object and object-memory representations across the proximodistal axis of CA1.

Episodic memory requires information about objects to be integrated into a spatial framework. Place cells in the hippocampus encode spatial representations of objects that could be generated through signaling from the entorhinal cortex. Projections from lateral (LEC) and medial entorhinal cortex (MEC) to the hippocampus terminate in distal and proximal CA1, respectively. We recorded place cells in distal and proximal CA1 as rats explored an environment that contained objects. Place cells in distal CA1 demonstrated higher measures of spatial tuning, stability, and closer proximity of place fields to objects. Furthermore, remapping to object displacement was modulated by place field proximity to objects in distal, but not proximal CA1. Finally, representations of previous object locations were closer to those locations in distal CA1 than proximal CA1. Our data suggest that in cue-rich environments, LEC inputs to the hippocampus support spatial representations with higher spatial tuning, closer proximity to objects, and greater stability than those receiving inputs from MEC. This is consistent with functional segregation in the entorhinal-hippocampal circuits underlying object-place memory.

Involvement of Hippocampal D1-Like Dopamine Receptors in the Inhibitory Effect of Cannabidiol on Acquisition and Expression of Methamphetamine-Induced Conditioned Place Preference.

Cannabidiol (CBD) is a non-psychotomimetic compound with strong potential to decrease the psychostimulant's rewarding effect with unclear receptors. Furthermore, as a part of the reward circuit, the hippocampus plays a crucial role in regulating the reward properties of drugs as determined by conditioned place preference (CPP). In the current research, CPP was used to evaluate the role of intra-CA1 microinjection of D1-like dopamine receptor antagonists in CBD's inhibitory effect on the acquisition and expression phases of methamphetamine (METH). Animals were treated by METH (1 mg/kg; sc) in a five-day schedule to induce CPP. To find out the impact of D1-like dopamine receptor antagonist, SCH23390, in the CA1 on the inhibitory influence of CBD on the acquisition of METH, the rats received intra-CA1 administration of SCH23390 (0.25, 1, and 4 µg/0.5 µl) following ICV treatment of CBD (10 µg/5 µl) over conditioning phase of METH. Furthermore, animals were given SCH23390 in the CA1 ensuing ICV microinjection of CBD (50 µg/5 µl) in the expression phase of METH to rule out the influence of SCH23390 on the suppressive effect of CBD on the expression of METH CPP. Intra-CA1 microinjection of SCH23390 abolished CBD's suppressive impact on both METH-induced CPP phases without any side effect on the locomotion. The current research disclosed that CBD inhibited the rewarding characteristic of METH via D1-like dopamine receptors in the CA1 region of the hippocampus.

Different effects of monophasic pulses and biphasic pulses applied by a bipolar stimulation electrode in the rat hippocampal CA1 region.

BACKGROUND: Electrical pulse stimulations have been applied in brain for treating certain diseases such as movement disorders. High-frequency stimulations (HFS) of biphasic pulses have been used in clinic stimulations, such as deep brain stimulation (DBS), to minimize the risk of tissue damages caused by the electrical stimulations. However, HFS sequences of monophasic pulses have often been used in animal experiments for studying neuronal responses to the stimulations. It is not clear yet what the differences of the neuronal responses to the HFS of monophasic pulses from the HFS of biphasic pulses are. METHODS: To investigate the neuronal responses to the two types of pulses, orthodromic-HFS (O-HFS) and antidromic-HFS (A-HFS) of biphasic and monophasic pulses (1-min) were delivered by bipolar electrodes, respectively, to the Schaffer collaterals (i.e., afferent fibers) and the alveus fibers (i.e., efferent fibers) of the rat hippocampal CA1 region in vivo. Evoked population spikes of CA1 pyramidal neurons to the HFSs were recorded in the CA1 region. In addition, single pulses of antidromic- and orthodromic-test stimuli were applied before and after HFSs to evaluate the baseline and the recovery of neuronal activity, respectively. RESULTS: Spreading depression (SD) appeared during sequences of 200-Hz monophasic O-HFS with a high incidence (4/5), but did not appear during corresponding 200-Hz biphasic O-HFS (0/6). A preceding burst of population spikes appeared before the SD waveforms. Then, the SD propagated slowly, silenced neuronal firing temporarily and resulted in partial recovery of orthodromically evoked population spikes (OPS) after the end of O-HFS. No SD events appeared during the O-HFS with a lower frequency of 100 Hz of monophasic or biphasic pulses (0/5 and 0/6, respectively), neither during the A-HFS of 200-Hz pulses (0/9). The antidromically evoked population spikes (APS) after 200-Hz biphasic A-HFS recovered to baseline level within ~ 2 min. However, the APS only recovered partially after the 200-Hz A-HFS of monophasic pulses. CONCLUSIONS: The O-HFS with a higher frequency of monophasic pulses can induce the abnormal neuron activity of SD and the A-HFS of monophasic pulses can cause a persisting attenuation of neuronal excitability, indicating neuronal damages caused by monophasic stimulations in brain tissues. The results provide guidance for proper stimulation protocols in clinic and animal experiments.

Increased Calbindin D28k Expression via Long-Term Alternate-Day Fasting Does Not Protect against Ischemia-Reperfusion Injury: A Focus on Delayed Neuronal Death, Gliosis and Immunoglobulin G Leakage.

Calbindin-D28k (CB), a calcium-binding protein, mediates diverse neuronal functions. In this study, adult gerbils were fed a normal diet (ND) or exposed to intermittent fasting (IF) for three months, and were randomly assigned to sham or ischemia operated groups. Ischemic injury was induced by transient forebrain ischemia for 5 min. Short-term memory was examined via passive avoidance test. CB expression was investigated in the Cornu Ammonis 1 (CA1) region of the hippocampus via western blot analysis and immunohistochemistry. Finally, histological analysis was used to assess neuroprotection and gliosis (microgliosis and astrogliosis) in the CA1 region. Short-term memory did not vary significantly between ischemic gerbils with IF and those exposed to ND. CB expression was increased significantly in the CA1 pyramidal neurons of ischemic gerbils with IF compared with that of gerbils fed ND. However, the CB expression was significantly decreased in ischemic gerbils with IF, similarly to that of ischemic gerbils exposed to ND. The CA1 pyramidal neurons were not protected from ischemic injury in both groups, and gliosis (astrogliosis and microgliosis) was gradually increased with time after ischemia. In addition, immunoglobulin G was leaked into the CA1 parenchyma from blood vessels and gradually increased with time after ischemic insult in both groups. Taken together, our study suggests that IF for three months increases CB expression in hippocampal CA1 pyramidal neurons; however, the CA1 pyramidal neurons are not protected from transient forebrain ischemia. This failure in neuroprotection may be attributed to disruption of the blood-brain barrier, which triggers gliosis after ischemic insults.

Interval-dependent neurotoxicity after multiple ketamine injections in late postnatal mice.

PURPOSE: Measuring the neurotoxic effects of multiple anesthetic exposures during neurodevelopment is complex due to the numerous factors that can affect the outcome. While we recently discovered that the interval between multiple sevoflurane exposures can affect the level of neurotoxicity, the significance of interval for other anesthetic agents is unknown. Thus, we evaluated the significance of dosing interval in the neurotoxic effects of multiple ketamine injections in postnatal day (PND) 17 mice. METHODS: PND17 mice of both sexes were intraperitoneally injected with ketamine (35 mg/kg) three times at short (2 h) or long (24 h) intervals. Changes in synaptic transmission were measured in hippocampal pyramidal neurons 5 days after the last injection, and behavioral changes were assessed at the age of 8 weeks. Values are presented as mean ± SD. RESULTS: Whereas short-interval ketamine injections enhanced excitatory synaptic transmission, as evidenced by an increased frequency of miniature excitatory postsynaptic currents (mEPSCs; ketamine, 0.09 ± 0.07 Hz; control, 0.06 ± 0.03 Hz), long-interval ketamine injections did not; instead, they decreased the amplitude of miniature inhibitory postsynaptic currents (mIPSCs; ketamine, 47.72 ± 6.90 pA; control, 51.21 ± 7.65 pA,). However, only long-interval ketamine injections induced long-term changes in anxiety behavioral in the open-field test (decrease in center duration; ketamine, 400.1 ± 162.8 s; control, 613.3 ± 312.7 s). CONCLUSIONS: Multiple ketamine injections induce interval-dependent, long-lasting synaptic changes and behavioral impairments. Future studies should carefully consider the dosing interval as a significant factor when studying the neurotoxic effects of multiple anesthetic exposures.

A novel method using ambient glutamate for the electrophysiological quantification of extrasynaptic NMDA receptor function in acute brain slices.

KEY POINTS: We present a novel protocol to quantify extrasynaptic NMDA receptor function utilizing the semi-selective activation of extrasynaptic receptors by ambient extracellular glutamate in acute brain slices from adult rats. We use whole cell patch clamp to measure the effect of the NMDA receptor antagonist MK-801 on both synaptic and brief, local agonist application-evoked responses. The level of ambient glutamate was estimated from tonic NMDA receptor activity to be ∼77 nM and an equivalent concentration of NMDA was used to estimate the degree of extrasynaptic blockade (>82%) by our MK-801 protocol. The extrasynaptic component of the total NMDA receptor pool can be mathematically derived from these data and was estimated to be 29-39% in the stratum radiatum of the CA1 region of the rat hippocampus. This technique could be used to quantify extrasynaptic NMDA receptor function in rodent models of diseases where extrasynaptic NMDA receptors are implicated in neuron death. ABSTRACT: Synaptic NMDA receptors (NMDARs) play a central role in pro-survival signalling and synaptic plasticity in the majority of excitatory synapses in the central nervous system whereas extrasynaptic NMDARs (ES-NMDARs) activate pro-death pathways and have been implicated in many neurodegenerative diseases. ES-NMDARs have been characterized in acute brain slice preparations using the largely irreversible, activity-dependent NMDAR antagonist MK-801 to block synaptic NMDARs. This approach is limited by the concomitant MK-801 blockade of ES-NMDARs activated by ambient extracellular glutamate, which is largely absent from the synaptic cleft due to the high density of nearby glutamate transporters. In acute hippocampal slices from rats aged 35-42 postnatal days, we estimated ambient glutamate to be 72-83 nM resulting in a block of more than 82% of ES-NMDARs during a 5 min MK-801 application. This paper describes a novel electrophysiological and mathematical method to quantify the proportion of NMDARs located at extrasynaptic locations in a confined region of an acute brain slice preparation using MK-801 to preferentially block ES-NMDARs. The protocol uses whole cell patch clamp measurement of NMDAR responses to synaptic stimulation and brief local pressure application of NMDA before and after MK-801 application. After mathematically correcting for the relative block of both synaptic and extrasynaptic receptors, ES-NMDARs were estimated to comprise 29-39% of the total NMDAR pool in the apical dendrites of hippocampal CA1 pyramidal neurons. This new method may prove useful for accurate quantification of NMDAR distributions in neurodegenerative diseases that are associated with increased toxic ES-NMDAR signalling.

Selective modulation of neuronal firing by pulse stimulations with different frequencies in rat hippocampus.

BACKGROUND: Deep brain stimulation (DBS) has a good prospect for treating many brain diseases. Recent studies have shown that axonal activation induced by pulse stimulations may play an important role in DBS therapies through wide projections of axonal fibers. However, it is undetermined whether the downstream neurons are inhibited or excited by axonal stimulation. The present study addressed the question in rat hippocampus by in vivo experiments. METHODS: Pulse stimulations with different frequencies (10-400 Hz) were applied to the Schaffer collateral, the afferent fiber of hippocampal CA1 region in anaesthetized rats. Single-unit spikes of interneurons and pyramidal cells in the downstream region of stimulation were recorded and evaluated. RESULTS: Stimulations with a lower frequency (10 or 20 Hz) did not change the firing rates of interneurons but decreased the firing rates of pyramidal cells (the principal neurons) significantly. The phase-locked firing of interneurons during these stimulations might increase the efficacy of GABAergic inhibitions on the principal neurons. However, stimulations with a higher frequency (100-400 Hz) increased the firing rates of both types of the neurons significantly. In addition, the increases of interneurons' firing were greater than the increases of pyramidal cells. Presumably, increase of direct excitation from afferent impulses together with failure of GABAergic inhibition might result in the increase of pyramidal cells' firing by a higher stimulation frequency. Furthermore, silent periods appeared immediately following the cessation of stimulations, indicating a full control of the neuronal firing by the stimulation pulses during axonal stimulation. Furthermore longer silent periods were associated with higher stimulation frequencies. CONCLUSIONS: Low-frequency (10-20 Hz) and high-frequency (100-400 Hz) stimulations of afferent axonal fibers exerted opposite effects on principal neurons in rat hippocampus CA1. These results provide new information for advancing deep brain stimulation to treat different brain disorders.

Hyper-response to Novelty Increases c-Fos Expression in the Hippocampus and Prefrontal Cortex in a Rat Model of Schizophrenia.

Schizophrenia is a debilitating disorder that may have a neurodevelopmental origin. For this reason, animal models based on neonatal insults or manipulations have been extensively used to demonstrate schizophrenia-related behaviors. Among those, the neonatal ventral hippocampus lesion (nVHL) is largely used as a model of schizophrenia-related behavior as it mimics behavioral and neurochemical abnormalities often seen in schizophrenic patients including hyperlocomotion in a novel environment. To investigate the neuroanatomical basis of coding novelty in the nVHL rat, we assessed the behavioral locomotor activity paradigm in a novel environment and measured expression of c-Fos, a marker of neural activation, in brain regions involved in the process of coding novelty or locomotion. Upon reaching adulthood, nVHL rats showed hyperlocomotion in the novel environment paradigm. Moreover, in nVHL rats the expression of c-Fos was greater in the prefrontal cortex (PFC) and CA1 region of the dorsal hippocampus compared to sham rats. Whereas similar expression of c-Fos was observed in the basolateral amygdala, nucleus accumbens and dentate gyrus region of  hippocampus of nVHL and sham rats. These results suggest that the nVHL disrupts the neural activity in the PFC and CA1 region of hippocampus in the process of coding novelty in the rat.

Comparison of the Role of D1- and D2-Like Receptors in the CA1 Region of the Hippocampus in the Reinstatement Induced by a Subthreshold Dose of Morphine and Forced Swim Stress in Extinguished Morphine-CPP in Rats.

Reward-seeking and relapse to drug use are two characteristics of addiction and reports have indicated the role of hippocampal structures in reward learning. To find the best ways of treatment, the understanding of the neurobiological mechanisms of reward and its involved factors is a must. For this reason, in the present study, we aimed to investigate the role of D1- and D2-like dopamine receptors and compared their activities in the CA1 region, focusing on the reinstatement induced by forced swim stress (FSS) or the combination of FSS and a subthreshold dose of morphine in extinguished morphine-CPP in rats. The rats were bilaterally implanted by two separate cannulas into the CA1 region. The animals received different doses of SCH23390 or sulpiride (0.5, 2, and 4 µg/0.5 µl vehicle/side) into the CA1 region on the reinstatement day and were tested for FSS-induced reinstatement or the combination of FSS and a subthreshold dose of morphine in separate groups. Our findings indicated that the D1- and D2-like receptor antagonists attenuated the reinstatement induced by the combination of FSS and the subthreshold dose of morphine. The behavioral results were more prominent in the groups of animals that received SCH23390 as compared to sulpiride. The data may suggest a role for the dopamine receptors in the CA1 region in relapse to drugs of abuse, which may be induced by exposure to a stressor.

Time-Course Change of Redd1 Expressions in the Hippocampal CA1 Region Following Chronic Cerebral Hypoperfusion.

Redd1, also known as RTP801/Dig2/DDIT4, is a stress-induced protein and marked changes of Redd1 expression occurs in response to hypoxia or cerebral ischemia. In the present study, we examined the time-course changes in Redd1 protein expressions in the rat hippocampal CA1 region following chronic cerebral hypoperfusion (CCH) induced by permanent bilateral common carotid arteries occlusion (2VO). Redd1 immunoreactivity in the pyramidal neurons of the hippocampal CA1 region was increased at 7 days after 2VO surgery, and then the immunoreactivity was decreased with time. Especially, very weak Redd1 immunoreactivity was observed in the hippocampal CA1 region at 28 days after 2VO surgery. Western blot analysis showed that Redd1 level in the hippocampal CA1 region was significantly increased at 7 days following CCH and significantly decreased at 28 days after 2VO surgery, compared with that of the sham-operated group. These results indicate that Redd1 expressions is markedly changed in the hippocampal CA1 region following CCH and that change of Redd1 expression may be associated with the CCH-induced neuronal damage in the hippocampal CA1 region.

Ten gigahertz microwave radiation impairs spatial memory, enzymes activity, and histopathology of developing mice brain.

For decades, there has been an increasing concern about the potential hazards of non-ionizing electromagnetic fields that are present in the environment and alarming as a major pollutant or electro-pollutant for health risk and neuronal diseases. Therefore, the objective of the present study was to explore the effects of 10 GHz microwave radiation on developing mice brain. Two weeks old mice were selected and divided into two groups (i) sham-exposed and (ii) microwave-exposed groups. Animals were exposed for 2 h/day for 15 consecutive days. After the completion of exposure, within an hour, half of the animals were autopsied immediately and others were allowed to attain 6 weeks of age for the follow-up study. Thereafter results were recorded in terms of various biochemical, behavioral, and histopathological parameters. Body weight result showed significant changes immediately after treatment, whereas non-significant changes were observed in mice attaining 6 weeks of age. Several other endpoints like brain weight, lipid peroxidation, glutathione, protein, catalase, and superoxide dismutase were also found significantly (p < 0.05) altered in mice whole brain. These significant differences were found immediately after exposure and also in follow-up on attaining 6 weeks of age in microwave exposure group. Moreover, statistically significant (p < 0.001) effect was investigated in spatial memory of the animals, in learning to locate the position of platform in Morris water maze test. Although in probe trial test, sham-exposed animals spent more time in searching for platform into the target quadrant than in opposite or other quadrants. Significant alteration in histopathological parameters (qualitative and quantitative) was also observed in CA1 region of the hippocampus, cerebral cortex, and ansiform lobule of cerebellum. Results from the present study concludes that the brain of 2 weeks aged mice was very sensitive to microwave exposure as observed immediately after exposure and during follow-up study at 6 weeks of age.

EARLY gestational exposure to isoflurane causes persistent cell loss in the dentate gyrus of adult male rats.

BACKGROUND: Our previous research showed that 4 h of maternal anesthesia with isoflurane during early gestation in pregnant rats leads to a deficit in spatial memory of adult male offspring. Because spatial memory is predominantly a hippocampally-mediated task, we asked the question if early gestational exposure to isoflurane affects development of the hippocampus in the offspring. FINDINGS: Previously behaviorally characterized adult male rats that were exposed to isoflurane during second trimester were sacrificed at 4 months of age (N = 10 and 13, control and isoflurane groups, respectively) for quantitative histology of hippocampal subregions. Sections were stained with cresyl violet and the total number of cells in the granular layer of the dentate gyrus and the pyramidal cell layer in the CA1 region were determined by a blinded observer using unbiased stereological principles and the optical fractionator method. Data were analyzed using Student's t test; P < 0.05 was accorded statistical significance. Stereological examination revealed 9% fewer cells in the granular layer of the dentate gyrus of isoflurane-exposed adult rats compared to controls (1,002,122 ± 84,870 vs. 1,091,829 ± 65,791, respectively; Mean ± S.D, *P = 0.01). In contrast, there were no changes in the cell number in the CA1 region, nor were there changes in the volumes of both regions. CONCLUSIONS: Our results show that maternal isoflurane anesthesia in rodents causes region-specific cell loss in the hippocampus of adult male offspring. These changes may, in part, account for the behavioral deficits reported in adult rats exposed to isoflurane in utero.

Cell-Permeable Peptide Targeting the Nrf2-Keap1 Interaction: A Potential Novel Therapy for Global Cerebral Ischemia.

The current study examined efficacy of a small Tat (trans-activator of transcription)-conjugated peptide activator of the Nrf2 (nuclear factor-E2-related factor-2) antioxidant/cell-defense pathway as a potential injury-specific, novel neuroprotectant against global cerebral ischemia (GCI). A competitive peptide, DEETGE-CAL-Tat, was designed to facilitate Nrf2 activation by disrupting interaction of Nrf2 with Keap1 (kelch-like ECH-associated protein 1), a protein that sequesters Nrf2 in the cytoplasm and thereby inactivates it. The DEETGE-CAL-Tat peptide contained the critical sequence DEETGE for the Nrf2-Keap1 interaction, the cell transduction domain of the HIV-Tat protein, and the cleavage sequence of calpain, which is sensitive to Ca(2+) increase and allows injury-specific activation of Nrf2. Using an animal model of GCI, we demonstrated that pretreatment with the DEETGE-CAL-Tat peptide markedly decreased Nrf2 interaction with Keap1 in the rat hippocampal CA1 region after GCI, and enhanced Nrf2 nuclear translocation and DNA binding. The DEETGE-CAL-Tat peptide also induced Nrf2 antioxidant/cytoprotective target genes, reduced oxidative stress, and induced strong neuroprotection and marked preservation of hippocampal-dependent cognitive function after GCI. These effects were specific as control peptides lacked neuroprotective ability. Intriguingly, the DEETGE-CAL-Tat peptide effects were also injury specific, as it had no effect upon neuronal survival or cognitive performance in sham nonischemic animals. Of significant interest, peripheral, postischemia administration of the DEETGE-CAL-Tat peptide from days 1-9 after GCI also induced robust neuroprotection and strongly preserved hippocampal-dependent cognitive function. Based on its robust neuroprotective and cognitive-preserving effects, and its unique injury-specific activation properties, the DEETGE-CAL-Tat peptide represents a novel, and potentially promising new therapeutic modality for the treatment of GCI. SIGNIFICANCE STATEMENT: The current study demonstrates that DEETGE-CAL-Tat, a novel peptide activator of a key antioxidant gene transcription pathway in the hippocampus after global cerebral ischemia, can exert robust neuroprotection and preservation of cognitive function. A unique feature of the peptide is that its beneficial effects are injury specific. This feature is attractive as it targets drug activation specifically in the site of injury, and likely would lead to a reduction of undesirable side effects if translatable to the clinic. Due to its injury-specific activation, robust neuroprotection, and cognitive-preserving effects, this novel peptide may represent a much-needed therapeutic advance that could have efficacy in the treatment of global cerebral ischemia.

Tea polyphenols may attenuate the neurocognitive impairment caused by global cerebral ischemia/reperfusion injury via anti-apoptosis.

BACKGROUND/AIMS: Global cerebral ischemia/reperfusion (GCIR) may incur neurocognitive impairment. Tea polyphenols (TP) have strong anti-oxidant capacity. This study planned to investigate the protective effect of TP against the neurocognitive impairment caused by GCIR and its mechanism. METHODS: One-stage anterior approach for cerebral four-vessel occlusion (4VO) was used to construct the GCIR model. Sprague Dawley rats were randomly classified into Sham group, GCIR group, and TP group (n = 50 per group). Besides receiving the same 4VO, the rats in TP group were treated with TP (6.4%) injection from the tail vein 30 minutes before cerebral ischemia. Morris water-maze test was used to evaluate the changes in space recognition and memory and open field activity test to assess the activity and motor function of rats. The cell apoptotic study in hippocampal CA1 region at specified time points (12, 24, 48, and 72 hours after surgery) was carried out by the flow cytometry, histology (hematoxylin and eosin staining), and immunohistochemical (terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining) examinations. One-way analysis of variance and least significant difference t-test were used and statistical significance considered at P < 0.05. RESULTS: Compared with the GCIR group, the TP group was significantly attenuated in the impairment of space recognition and memory caused by GCIR and so was the neuronal apoptosis in the hippocampal CA1 region (P < 0.05). CONCLUSION: TP may attenuate the impairment of space recognition and memory caused by GCIR via anti-apoptosis.

Early postnatal nicotine exposure causes hippocampus-dependent memory impairments in adolescent mice: Association with altered nicotinic cholinergic modulation of LTP, but not impaired LTP.

Fetal nicotine exposure from smoking during pregnancy causes long-lasting cognitive impairments in offspring, yet little is known about the mechanisms that underlie this effect. Here we demonstrate that early postnatal exposure of mouse pups to nicotine via maternal milk impairs long-term, but not short-term, hippocampus-dependent memory during adolescence. At the Schaffer collateral (SC) pathway, the most widely studied synapses for a cellular correlate of hippocampus-dependent memory, the induction of N-methyl-D-aspartate receptor-dependent transient long-term potentiation (LTP) and protein synthesis-dependent long-lasting LTP are not diminished by nicotine exposure, but rather unexpectedly the threshold for LTP induction becomes lower after nicotine treatment. Using voltage sensitive dye to visualize hippocampal activity, we found that early postnatal nicotine exposure also results in enhanced CA1 depolarization and hyperpolarization after SC stimulation. Furthermore, we show that postnatal nicotine exposure induces pervasive changes to the nicotinic modulation of CA1 activity: activation of nicotinic receptors no longer increases CA1 network depolarization, acute nicotine inhibits rather than facilitates the induction of LTP at the SC pathway by recruiting an additional nicotinic receptor subtype, and acute nicotine no longer blocks LTP induction at the temporoammonic pathway. These findings reflect the pervasive impact of nicotine exposure during hippocampal development, and demonstrate an association of hippocampal memory impairments with altered nicotinic cholinergic modulation of LTP, but not impaired LTP. The implication of our results is that nicotinic cholinergic-dependent plasticity is required for long-term memory formation and that postnatal nicotine exposure disrupts this form of plasticity.

Rhynchophylline Protects Against the Amyloid ß-Induced Increase of Spontaneous Discharges in the Hippocampal CA1 Region of Rats.

Accumulated soluble amyloid ß (Aß)-induced aberrant neuronal network activity has been recognized as a key causative factor leading to cognitive deficits which are the most outstanding characteristic of Alzheimer's disease (AD). As an important structure associated with learning and memory, the hippocampus is one of the brain regions that are impaired very early in AD, and the hippocampal CA1 region is selectively vulnerable to soluble Aß oligomers. Our recent study showed that soluble Aß1-42 oligomers induced hyperactivity and perturbed the firing patterns in hippocampal neurons. Rhynchophylline (RIN) is an important active tetracyclic oxindole alkaloid isolated from Uncaria rhynchophylla which is a traditional Chinese medicine and often used to treat central nervous system illnesses such as hypertension, convulsions, tremor, stroke etc. Previous evidence showed that RIN possessed neuroprotective effects of improving the cognitive function of mice with Alzheimer-like symptoms. In the present study, we aimed to investigate the protective effect of RIN against soluble Aß1-42 oligomers-induced hippocampal hyperactivity. The results showed that (1) the mean frequency of spontaneous discharge was increased by the local application of 3 µM soluble Aß1-42 oligomers; (2) 30 µM RIN did not exert any obvious effects on basal physiological discharges; and (3) treatment with RIN effectively inhibited the soluble Aß1-42 oligomers-induced enhancement of spontaneous discharge, in a concentration-dependent manner with an IC50 = 9.0 µM. These in vivo electrophysiological results indicate that RIN can remold the spontaneous discharges disturbed by Aß and counteract the deleterious effect of Aß1-42 on neural circuit. The experimental findings provide further evidence to affirm the potential of RIN as a worthy candidate for further development into a therapeutic agent for AD.

Developmental increase in hippocampal endocannabinoid mobilization: role of metabotropic glutamate receptor subtype 5 and phospholipase C.

Endocannabinoids (eCBs) released from postsynaptic neurons mediate retrograde suppression of neurotransmitter release at central synapses. eCBs are crucial for establishing proper synaptic connectivity in the developing nervous system. Mobilization of eCBs is driven either by a rise in intracellular Ca(2+) (depolarization-induced suppression of inhibition, DSI) or postsynaptic G protein-coupled receptors (GPCRs) that activate phospholipase C beta (PLCß). To determine whether eCB mobilization changes between neonatal and juvenile ages, we used whole cell voltage-clamp recordings of CA1 neurons from rat hippocampal slices at postnatal days 1-18 (neonatal) and 19-43 (juvenile), because many neurophysiological parameters change dramatically between approximately postnatal days 18-20. We found that DSI was slightly greater in juveniles than in neonates, while eCB mobilization stimulated by GPCRs was unchanged. However, when DSI was elicited during GPCR activation, its increase was much greater in juveniles, suggesting that eCB mobilization caused by the synergy between the Ca(2+) and GPCR pathways is developmentally upregulated. Western blotting revealed significant increases in both metabotropic type glutamate receptor 5 (mGluR5) and PLCß1 proteins in juveniles compared with neonates. Responses to pharmacological activation or inhibition of PLC implied that eCB upregulation is associated with a functional increase in PLC activity. We conclude that synergistic eCB mobilization in hippocampal CA1 neurons is greater in juveniles than in neonates, and that this may result from increases in the mGluR5-PLCß1 eCB pathway. The data enhance our understanding of the developmental regulation of the eCB system and may provide insight into diseases caused by improper cortical wiring, or the impact of cannabis exposure during development.