Yoshito Saitoh

Adult Neurogenesis Modulates the Hippocampus-Dependent Period of Associative Fear Memory

Acquired memory initially depends on the hippocampus (HPC) for the process of cortical permanent memory formation. The mechanisms through which memory becomes progressively independent from the HPC remain unknown. In the HPC, adult neurogenesis has been described in many mammalian species, even at old ages. Using two mouse models in which hippocampal neurogenesis is physically or genetically suppressed, we show that decreased neurogenesis is accompanied by a prolonged HPC-dependent period of associative fear memory. Inversely, enhanced neurogenesis by voluntary exercise sped up the decay rate of HPC dependency of memory, without loss of memory. Consistently, decreased neurogenesis facilitated the long-lasting maintenance of rat hippocampal long-term potentiation in vivo. These independent lines of evidence strongly suggest that the level of hippocampal neurogenesis play a role in determination of the HPC-dependent period of memory in adult rodents. These observations provide a framework for understanding the mechanisms of the hippocampal-cortical complementary learning systems.

Novel Members of the Vesl/Homer Family of PDZ Proteins That Bind Metabotropic Glutamate Receptors

Vesl-1S (186 amino acids, also called Homer) is a protein containing EVH1- and PDZ-like domains whose expression in the hippocampus is regulated during long term potentiation (LTP), one form of synaptic plasticity thought to underlie memory formation (Kato, A., Ozawa, F., Saitoh, Y., Hirai, K., and Inokuchi, K. (1997) FEBS Lett. 412, 183–189; Brakeman, P. R., Lanahan, A. A., O’Brien, R., Roche, K., Barnes, C. A., Huganir, R. L., and Worley, P. F. (1997)Nature 386, 284–288). Here we report additional members of the Vesl/Homer family of proteins, Vesl-1L and Vesl-2. Vesl-1L (366 amino acids), a splicing variant of Vesl-1S, shares N-terminal 175 amino acids with Vesl-1S and contains additional amino acids at the C terminus. Vesl-2 (354 amino acids) was highly related to Vesl-1L in that both contain EVH1- and PDZ-like domains at the N terminus (86% conservation) and an MCC (mutated in colorectal cancer)-like domain and a leucine zipper at the C terminus. In contrast to vesl-1S, we observed no changes in the levels of vesl-1L andvesl-2 mRNAs during dentate gyrus LTP. All these proteins interacted with metabotropic glutamate receptors (mGluR1 and mGluR5) as well as several hippocampal proteins in vitro. Vesl-1L and Vesl-2, but not Vesl-1S, interacted with each other through the C-terminal portion that was absent in Vesl-1S. Vesl-1L and Vesl-2 may mediate clustering of mGluRs at synaptic junctions. We propose that Vesl-1S may be involved in the structural changes that occur at metabotropic glutamatergic synapses during the maintenance phase of LTP by modulating the redistribution of synaptic components.

vesl, a gene encoding VASP/Ena family related protein, is upregulated during seizure, long-term potentiation and synaptogenesis.

We have isolated a novel cDNA, vesl, that was induced during convulsive seizure in the rat hippocampus. The vesl gene encodes a protein of 186 amino acids that has significant homology to the EVH1 domain of the VASP/Ena family of proteins implicated in the control of microfilament dynamics. The expression of vesl mRNA was induced in the granule cell layer during persistent long‐term potentiation (LTP) of the dentate gyrus in an NMDA receptor‐dependent manner. Furthermore, vesl mRNA was expressed at a high level during hippocampal synaptogenesis. We suggest that the Vesl protein may be involved in the structural changes that occur at synapses during long‐lasting neuronal plasticity and development.

Identification and Cataloging of Genes Induced by Long‐Lasting Long‐Term Potentiation in Awake Rats

Abstract: Maintenance of long‐term potentiation (LTP) requires de novo gene expression. Here we report the direct isolation, using PCR‐differential display, of genes whose expression level was altered after induction of long‐lasting LTP in the hippocampus of freely moving awake rats. Differential display using 480 primer combinations revealed 17 cDNA bands that showed a reproducible change in expression level. These cDNAs represented at least 10 different genes (termed RM1‐10), all of which showed up‐regulation at 75 min after LTP induction and a return to basal expression levels within 24 h. Three of these genes were known only from expressed sequence tags (RM1‐3), two were known genes whose up‐regulation by LTP has not been described (GADD153/CHOP and ler5), and five were known genes whose up‐regulation by LTP has already been reported (MAPK phosphatase, NGFI‐A/zif268, vesl‐1S/homer‐1a, Ag2, and krox‐20). We characterized the expression profiles of genes in the two former categories with respect to NMDA receptor dependency, tissue specificity, and developmental regulation using northern blotting and semiquantitative RT‐PCR. The up‐regulation of all five of these genes was NMDA receptor‐dependent and correlated with the persistence of LTP, suggesting that these genes may play functional roles in prolonged LTP maintenance.

LTP induction within a narrow critical period of immature stages enhances the survival of newly generated neurons in the adult rat dentate gyrus

Neurogenesis occurs in the adult hippocampus of various animal species. A substantial fraction of newly generated neurons die before they mature, and the survival rate of new neurons are regulated in an experience-dependent manner. Previous study showed that high-frequency stimulation (HFS) of perforant path fibers to the hippocampal dentate gyrus (DG) induces the long-term potentiation (LTP) in the DG, and enhances the survival of newly generated neurons in the DG. In this study, we addressed whether a time period exists during which the survival of new neurons is maximally sensitive to the HFS. We found that the enhancement of cell survival by HFS was exclusively restricted to the specific narrow period during immature stages of new neurons (7-10 days after birth). Furthermore, the pharmacological blockade of LTP induction suppressed the enhancement of cell survival by the HFS. These results suggest that the LTP induction within a narrow critical period of immature stages enhances the survival of newly generated neurons in rat DG.

Cross-species transplantation of the suprachiasmatic nuclei from rats to siberian chipmunks (Eutamias sibiricus) with suprachiasmatic lesions

Suprachiasmatic nuclei (SCN) obtained from neonatal or embryonic 19 or 20 day rats, were grafted into the third ventricle of SCN-lesioned arrhythmic siberian chipmunks. Four out of 37 chipmunks showed reappearance of circadian rhythmicity in wheel running activity. In all 4 cases, at least one surviving graft was confirmed in the host brain. Also, vasopressin and vasoactive intestinal polypeptide (VIP)-like immunoreactive substances were found in the graft, suggesting the existence of live SCN neurons. Although the number of successful cases and the intensity of the restored rhythm was limited compared to the intra-species grafting in rats, a possibility that cross-species transplantation of SCN can restore circadian rhythmicity was shown.

Location of the suprachiasmatic nucleus grafts in rats which restored circadian rhythmicity after transplantation

In Wistar male rats whose circadian wheel running activity rhythms were disrupted by bilateral suprachiasmatic nuclei (SCN) lesions, transplantation of neonatal rat SCN into the 3rd ventricle was performed. Out of 49 rats from which adequate wheel running activity records were obtained, 15 rats showed restoration of circadian rhythm starting 2 to 13 weeks (average 1 month) after transplantation. The existence of active SCN neurons in the graft was shown by immunocytochemical reactivity to vasopressin- and VIP-like substances. In all rats, effective grafts were found in the diencephalon, mostly on the wall of the 3rd ventricle.

Activity-dependent localization in spines of the F-actin capping protein CapZ screened in a rat model of dementia

Actin reorganization in dendritic spines is hypothesized to underlie neuronal plasticity. Actin‐related proteins, therefore, might serve as useful markers of plastic changes in dendritic spines. Here, we utilized memory deficits induced by fimbria‐fornix transection (FFT) in rats as a dementia model to screen candidate memory‐associated molecules by using a two‐dimensional gel method. Comparison of protein profiles between the transected and control sides of hippocampi after unilateral FFT revealed a reduction in the F‐actin capping protein (CapZ) signal on the FFT side. Subsequent immunostaining of brain sections and cultured hippocampal neurons revealed that CapZ localized in dendritic spines and the signal intensity in each spine varied widely. The CapZ content decreased after suppression of neuronal firing by tetrodotoxin treatment in cultured neurons, indicating rapid and activity‐dependent regulation of CapZ accumulation in spines. To test input specificity of CapZ accumulation in vivo, we delivered high‐frequency stimuli to the medial perforant path unilaterally in awake rats. This path selectively inputs to the middle molecular layer of the dentate gyrus, where CapZ immunoreactivity increased. We conclude that activity‐dependent, synapse‐specific regulation of CapZ redistribution might be important in both maintenance and remodeling of synaptic connections in neurons receiving specific spatial and temporal patterns of inputs.

Spine Formation Pattern of Adult-Born Neurons Is Differentially Modulated by the Induction Timing and Location of Hippocampal Plasticity

In the adult hippocampus dentate gyrus (DG), newly born neurons are functionally integrated into existing circuits and play important roles in hippocampus-dependent memory. However, it remains unclear how neural plasticity regulates the integration pattern of new neurons into preexisting circuits. Because dendritic spines are major postsynaptic sites for excitatory inputs, spines of new neurons were visualized by retrovirus-mediated labeling to evaluate integration. Long-term potentiation (LTP) was induced at 12, 16, or 21 days postinfection (dpi), at which time new neurons have no, few, or many spines, respectively. The spine expression patterns were investigated at one or two weeks after LTP induction. Induction at 12 dpi increased later spinogenesis, although the new neurons at 12 dpi didn’t respond to the stimulus for LTP induction. Induction at 21 dpi transiently mediated spine enlargement. Surprisingly, LTP induction at 16 dpi reduced the spine density of new neurons. All LTP-mediated changes specifically appeared within the LTP–induced layer. Therefore, neural plasticity differentially regulates the integration of new neurons into the activated circuit, dependent on their developmental stage. Consequently, new neurons at different developmental stages may play distinct roles in processing the acquired information by modulating the connectivity of activated circuits via their integration.

A triphasic curve characterizes the retention of lever-pressing behavior rewarded by lateral hypothalamic stimulation during the immediate-post-trial period in rats: implications for a transient-intermediate stage between short-and long-term memory

We investigated the retention of lever-pressing behavior in rats rewarded by lateral hypothalamic (LHT) stimulation. Rats with electrodes implanted into the lateral hypothalamus were trained to press a lever to receive LHT stimulation. More than 90% of rats which had self-stimulated for longer than 30 min retained this behavior when tested 1 or 7 days later, indicating that intracranial self-stimulation (ICSS) produced a stable long-term memory lasting for at least 7 days. Subsequently, we examined retention at shorter ICSS-test intervals. Retention rates were measured 15, 30, 60, 90, 120, 180 or 360 min after the end of a 90-min period of ICSS, using a different group of 15 rats for each interval. Unexpectedly, retention rates in shorter interval tests were lower than those observed after 1 or 7 days. We observed a characteristic fluctuation in retention rates with lower rates at 30 and 180 min, giving a triphasic form to the retention curve with peaks at 15, 90 and 360 min. When a group of rats that had been previously stimulated and shown to have retained the lever-pressing behavior was allowed to re-stimulate a second time, no fluctuations were observed in short-term interval tests. This indicates that the fluctuations in the retention curve immediately after initial ICSS are closely related to the initial acquisition of the memory. Our results support a three-phase model of memory formation that includes a transient-intermediate stage between short- and long-term memory.