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Summary hypothesis is that an excess supply of target-derived neurotrophins should eliminate competition among thalMaturation of the visual cortex is influenced by visual amic inputs and therefore prevent the formation of ocuexperience during an early postnatal period. The fac- lar dominance columns or the physiological shift of ocutors that regulate such a critical period remain unclear. lar dominance following monocular deprivation (MD). We examined the maturation and plasticity of the vi- Indeed, intracortical administrations of BDNF or its neusual cortex in transgenic mice in which the postnatal tralizing agent, the trkB-IgG fusion protein, were shown rise of brain-derived neurotrophic factor (BDNF) was to alter the formation (Cabelli et al., 1995, 1997) and accelerated. In these mice, the maturation of GABAer- plasticity (Galuske et al., 1996) of ocular dominance colgic innervation and inhibition was accelerated. Fur- umns. Furthermore, cortical infusion of NT-4 rescued thermore, the age-dependent decline of cortical long- LGN neurons from the effect of MD (Riddle et al., 1995). term potentiation induced by white matter stimulation, These results are consistent with the notion that a ligand a form of synaptic plasticity sensitive to cortical inhibi- of TrkB, either BDNF and/or NT-4, acts directly on LGN tion, occurred earlier. Finally, transgenic mice showed axons and is involved in their activity-dependent compea precocious development of visual acuity and an ear- tition for layer IV synaptic targets during ocular domilier termination of the critical period for ocular domi- nance segregation. However, alternative interpretations nance plasticity. We propose that BDNF promotes the for these findings are plausible. For example, neuromaturation of cortical inhibition during early postnatal trophins could affect LGN axons indirectly, via alterlife, thereby regulating the critical period for visual ations in the dendritic morphology of cortical neurons, cortical plasticity.

Carbon monoxide (CO), generated in neurons by the enzyme heme oxygenase-2 (HO2), is postulated to be a gaseous signaling molecule in the mammalian brain. Because of the recent evidence suggesting a...

In this study, we compared the influence of two peptides on the selection of CD8αα and CD8αβ intraepithelial lymphocytes (IELs) of the intestine, which develop by a unique and partially thymus-independent process. Mice were used in which all T cells carried one transgenic T cell antigen receptor (TCR) (F5), and in which only well defined transgenic peptides were presented by H-2Db. The first peptide, for which the F5 TCR has a high affinity, derives from the influenza virus nucleoprotein (NP68). The second peptide, NP34, is an antagonistic variant of NP68 and is recognized by the F5 TCR with low affinity. To avoid presentation of endogenous peptides or production of T cells carrying alternative TCRs, F5 TCR transgenic mice were generated that were deficient for Tap-1 and Rag-1. In these mice, no CD3+CD8+cells were found in lymph nodes, spleen, or intestine. Introduction of transgenes encoding either NP34 or NP68 along with an endoplasmic reticulum signal sequence enabled Tap-1-independent expression of each peptide in these mice. Positive selection of F5TCR+CD8+thymocytes was not rescued by these transgenic peptides. However, the high-affinity NP68 peptide induced maturation of CD8αα IEL, whereas the low-affinity NP34 peptide stimulated development of both CD8αβ and CD8αα IEL, but in smaller numbers. When both peptides were present, CD8αβ T cells failed to develop and the number of CD8αα IELs was lower than in mice carrying the NP68 transgene alone. These data demonstrate that single ligands with a high or low affinity for TCR are capable of inducing or inhibiting the maturation of alternative subsets of IELs.

Multiple sclerosis (MS) is a severe central nervous system disease. Experimental autoimmune encephalomyelitis (EAE) mimics MS in mice. We report that spontaneous development of EAE in RAG-1-deficient mice transgenic for a myelin basic protein (MBP)-specific TCR (TgMBP+/RAG-1−/−) requires expression of the T cell costimulatory molecule CD28. Surprisingly, T cells from CD28−/−TgMBP+/RAG-1−/− mice proliferate and produce IL-2 in response to MBP1–17 peptide in vitro, excluding clonal anergy as the mechanism of CD28-regulated pathogenesis. Proliferation of autoaggressive T cells was dependent on the concentration of the MBP peptide, as was the development of MBP-induced EAE in CD28-deficient PL/J mice. These results provide the first genetic evidence that CD28 costimulation is crucial for MBP-specific T cell activation in vivo and the initiation of spontaneous EAE.

Previous studies using a variety of drugs with different affinities for the dopamine (DA) D3 receptor suggested that this receptor is involved in regulating motor activity and hypothermia. However, the in vivo selectivity of many of these compounds has been repeatedly questioned. To examine the precise roles of the DA D3 receptor in motor activity and hypothermic responses, we used mutant mice lacking the DA D3 receptor to evaluate the in vivo effects of several putative D3 receptor-selective agonists and antagonists. Using automated photocell activity chambers, we observed that the decreases in locomotor activity produced by putative D3 receptor-selective agonists as well as increases in locomotor activity produced by putative D3 receptor antagonists are identical in D3 receptor mutant and wild-type mice. In addition, the hypothermia produced by the putative D3 receptor-selective agonist PD 128907 is identical in both groups of mice. Based on these findings, we propose that D3 receptors are unlikely to be involved in these effects and we caution that the putative D3 ligands that have been used to reach conclusions regarding the functional roles of D3 receptors lack the necessary in vivo selectivity to support such conclusions.

Initial sensitivity and tolerance development to the sedative-hypnotic and hypothermic effects of ethanol were investigated in gamma-protein kinase C (PKC) null mutant mice. Null mutants from a C57BL/6J x 129/SvJ mixed genetic background demonstrated decreased ethanol sensitivity and failed to develop chronic tolerance after 10 days of ethanol liquid diet. However, when the null mutation was introgressed onto a C57BL/6J background for six generations, the "no tolerance" phenotype for sedative-hypnotic and hypothermic effects of ethanol was no longer apparent Outcrossing the gamma-PKC null mutation to a C57BL/6J x 129/SvEvTac mixed background restored the "no tolerance" phenotype to ethanol-induced sedation after chronic ethanol diet; however, as measured by hypothermia, tolerance was still evident in the null mutant mice. These observations and the results of tests of chronic tolerance in the C57BL/6J, 129/SvJ, and 129/SvEvTac background inbred strains indicate that gamma-PKC plays an important role in initial sensitivity and tolerance to ethanol. However, the impact of gamma-PKC is modulated by the background genotype. These results stress the importance of including the effect of genetic background when evaluating the effects of single gene mutations on quantitative behavioral traits.

Autoimmune diseases result from a failure of tolerance. Although many self-reactive T cells are present in animals and humans, their activation appears to be prevented normally by regulatory T cells. In this study, we show that regulatory CD4(+) T cells do protect mice against the spontaneous occurrence of experimental autoimmune encephalomyelitis (EAE), a mouse model for multiple sclerosis. Anti-myelin basic protein (MBP) TCR transgenic mice (T/R+) do not spontaneously develop EAE although many self-reactive T cells are present in their thymi and peripheral lymphoid organs. However, the disease develops in all crosses of T/R+ mice with recombination-activating gene (RAG)-1 knockout mice in which transgenic TCR-expressing cells are the only lymphocytes present (T/R- mice). In this study, crosses of T/R+ mice with mice deficient for B cells, CD8(+) T cells, NK1.1 CD4(+) T (NKT) cells, gamma/delta T cells, or alpha/beta T cells indicated that alpha/beta CD4(+) T cells were the only cell population capable of controlling the self-reactive T cells. To confirm the protective role of CD4(+) T cells, we performed adoptive transfer experiments. CD4(+) T cells purified from thymi or lymph nodes of normal mice prevented the occurrence of spontaneous EAE in T/R- mice. To achieve full protection, the cells had to be transferred before the recipient mice manifested any symptoms of the disease. Transfer of CD4(+) T cells after the appearance of symptoms of EAE had no protective effect. These results indicate that at least some CD4(+) T cells have a regulatory function that prevent the activation of self-reactive T cells.

Factors inside and outside neurons control the process of axonal growth and regeneration. Recently, it has become apparent that neurons are determined intrinsically for their ability to grow axons. In the mammalian CNS, the intrinsic machinery of neurons that triggers the growth of axons during early embryonic stages is shut down at a certain point in development; as a consequence, axon elongation and regeneration cannot occur in postnatal life. The proto-oncogene Bcl-2 has been recognized to act as a key regulator for the program of axon elongation inside neurons. However, expressing the gene Bcl-2 in CNS neurons is not sufficient to induce nerve regeneration in the adult CNS, eliminating the inhibitory mechanism in the mature CNS environment is still required. Recently, the formation of glia scar has been reported to be the major limiting factor in the CNS environment that blocks nerve regeneration. These new discoveries challenge the classical view of nerve regeneration in the mammalian CNS. It opens up a new dimension in the study of the cellular and molecular mechanisms underlying neurodevelopmental and neurodegenerative diseases.

Previous work has shown that mice missing the α-isoform of calcium–calmodulin-dependent protein kinase II (α-CaMKII) have a deficiency in CA1 hippocampal long-term potentiation (LTP). Follow-up studies on subsequent generations of these mutant mice in a novel inbred background by our laboratories have shown that whereas a deficiency in CA1 LTP is still present in α-CaMKII mutant mice, it is different both quantitatively and qualitatively from the deficiency first described. Mice of a mixed 129SvOla/SvJ;BALB/c;C57Bl/6 background derived from brother/sister mating of the α-CaMKII mutant line through multiple generations (>10) were produced by use of in vitro fertilization. Although LTP at 60 min post-tetanus was clearly deficient in these (−/−) α-CaMKII mice (42.6%, n = 33) compared with (+/+) α-CaMKII control animals (81.7%,n = 17), α-CaMKII mutant mice did show a significant level of LTP. The amount of LTP observed in α-CaMKII mutants was normally distributed, blocked by APV (2.7%, n = 8), and did not correlate with age. Although this supports a role for α-CaMKII in CA1 LTP, it also suggests that a form of α-CaMKII-independent LTP is present in mice that could be dependent on another kinase, such as the β-isoform of CaMKII. A significant difference in input/output curves was also observed between (−/−) α-CaMKII and (+/+) α-CaMKII animals, suggesting that differences in synaptic transmission may be contributing to the LTP deficit in mutant mice. However, tetani of increasing frequency (50, 100, and 200 Hz) did not reveal a higher threshold for potentiation in (−/−) α-CaMKII mice compared with (+/+) α-CaMKII controls.

Nitric oxide (NO) is well established as a neurotransmitter in the central and peripheral nervous systems. More recently, another gas, carbon monoxide (CO) has also been implicated in neurotransmission. In the nervous system CO is formed by a subtype of heme oxygenase (HO) designated HO2. HO2 is localized to discrete neuronal populations in the brain resembling localizations of soluble guanylyl cyclase, which is activated by CO. CO may also function in the peripheral autonomic nervous system, in conjunction with NO. The majority of ganglia in the myenteric plexus possess both HO2 and neuronal NO synthase (NOS). Defects in myenteric plexus neurotransmission occur both in mice with targeted deletion of genes for HO2 and neuronal NOS. HO2 also occurs in other autonomic ganglia including the petrosal, superior cervical and nodose ganglia. Neuronal NOS is localized to neurons regulating male reproductive behavior, such as penile erection, and NOS inhibitors prevent erection. Because of the other parallels between NO and CO, we speculated that CO may play a role in male reproductive behavior. In the present study we describe HO2 localization in neuronal structures regulating copulatory reflexes. Reflex activity of the bulbospongiosus muscle, which mediates ejaculation and ejaculatory behavior, is markedly diminished in mice with targeted deletion of the gene for HO2 (HO2-).

Neuronal nitric oxide synthase (nNOS) generates NO in neurons, and heme-oxygenase-2 (HO-2) synthesizes carbon monoxide (CO). We have evaluated the roles of NO and CO in intestinal neurotransmission using mice with targeted deletions of nNOS or HO-2. Immunohistochemical analysis demonstrated colocalization of nNOS and HO-2 in myenteric ganglia. Nonadrenergic noncholinergic relaxation and cyclic guanosine 3′,5′ monophosphate elevations evoked by electrical field stimulation were diminished markedly in both nNOS Δ / Δ and HO-2 Δ / Δ mice. In wild-type mice, NOS inhibitors and HO inhibitors partially inhibited nonadrenergic noncholinergic relaxation. In nNOS Δ / Δ animals, NOS inhibitors selectively lost their efficacy, and HO inhibitors were inactive in HO-2 Δ / Δ animals.

We have examined the role of NMDA receptor–mediated neural activity in the formation of periphery-related somatosensory patterns, using genetically engineered mice. We demonstrate that ectopic expression of a transgene of an NMDAR1 splice variant rescues neonatally fatal NMDAR1 knockout (KO) mice, although the average life span varies depending on the level of the transgene expression. In NMDAR1 KO mice with “high” levels of the transgene expression, sensory periphery-related patterns were normal along both the trigeminal and dorsal column pathways. In the KO mice with “low” levels of the transgene expression, the patterns were absent in the trigeminal pathway. Our results indicate that NMDA receptor–mediated neural activity plays a critical role in pattern formation along the ascending somatosensory pathways.

In normal animals, peripheral nerve injury produces a persistent, neuropathic pain state in which pain is exaggerated and can be produced by nonpainful stimuli. Here, mice that lack protein kinase C gamma (PKCγ) displayed normal responses to acute pain stimuli, but they almost completely failed to develop a neuropathic pain syndrome after partial sciatic nerve section, and the neurochemical changes that occurred in the spinal cord after nerve injury were blunted. Also, PKCγ was shown to be restricted to a small subset of dorsal horn neurons, thus identifying a potential biochemical target for the prevention and therapy of persistent pain.

Most of the gammadelta T cells in the intestinal epithelium of normal mice use the Vgamma1 or the Vgamma7 gene segments. However, the relative proportions of gammadelta intraepithelial lymphocytes expressing either the Vgamma1 or the Vgamma7 chain vary among different strains of mice whereas they are quite constant between different individuals of the same strain, suggesting that genetic factors, rather than environmental factors, are responsible for the observed differences. To analyze the genetic factors influencing the representation of different gammadelta T cell subsets in the intestinal epithelium, we used available anti-T cell antigen receptor (TCR) V region-specific mAbs against Vgamma1, Vgamma4, Vgamma7, and Vdelta4 to examine the TCR repertoire of intraepithelial gammadelta lymphocytes in a set of (C57BL/6 x DBA/2) recombinant inbred strains. Our results show that the representation of different Vgamma and Vdelta gene products among gammadelta intestinal intraepithelial lymphocytes is under a complex genetic control with a marked influence by genes closely linked to the TCRgamma, TCRdelta, and major histocompatibility complex loci.

Most of the cerebellar Purkinje cells (PCs) of an adult animal are innervated individually by a single climbing fiber (CF) that forms strong excitatory synapses with the PCs. This one-to-one relationship between a PC and a CF is a consequence of a developmentally regulated regression of the innervation of PCs by CFs. We found that, in mice deficient in the type 1 metabotropic glutamate receptor (mGluR1), the regression of supernumerary CFs ceases by the end of the second postnatal week, which is about one week earlier than in normal mice. Consequently, about one third of PCs in the mGluR1 mutant mice are innervated by multiple CFs in adulthood. We conclude that the regression of CFs normally occurs in two developmental phases and that mGluR1 plays a crucial role in the second phase.

The valence of memories is malleable because of their intrinsic reconstructive property. This property of memory has been used clinically to treat maladaptive behaviours. However, the neuronal mechanisms and brain circuits that enable the switching of the valence of memories remain largely unknown. Here we investigated these mechanisms by applying the recently developed memory engram cell- manipulation technique. We labelled with channelrhodopsin-2 (ChR2) a population of cells in either the dorsal dentate gyrus (DG) of the hippocampus or the basolateral complex of the amygdala (BLA) that were specifically activated during contextual fear or reward conditioning. Both groups of fear-conditioned mice displayed aversive light-dependent responses in an optogenetic place avoidance test, whereas both DG- and BLA-labelled mice that underwent reward conditioning exhibited an appetitive response in an optogenetic place preference test. Next, in an attempt to reverse the valence of memory within a subject, mice whose DG or BLA engram had initially been labelled by contextual fear or reward conditioning were subjected to a second conditioning of the opposite valence while their original DG or BLA engram was reactivated by blue light. Subsequent optogenetic place avoidance and preference tests revealed that although the DG-engram group displayed a response indicating a switch of the memory valence, the BLA-engram group did not. This switch was also evident at the cellular level by a change in functional connectivity between DG engram-bearing cells and BLA engram-bearing cells. Thus, we found that in the DG, the neurons carrying the memory engram of a given neutral context have plasticity such that the valence of a conditioned response evoked by their reactivation can be reversed by re-associating this contextual memory engram with a new unconditioned stimulus of an opposite valence. Our present work provides new insight into the functional neural circuits underlying the malleability of emotional memory., RIKEN Brain Science Institute, Howard Hughes Medical Institute, JPB Foundation, National Institutes of Health (U.S.) (Pre-doctoral Training Grant T32GM007287)

Defects in p21-activated kinase (PAK) are suspected to play a role in cognitive symptoms of Alzheimer's disease (AD). Dysfunction in PAK leads to cofilin activation, drebrin displacement from its actin-binding site, actin depolymerization/severing, and, ultimately, defects in spine dynamics and cognitive impairment in mice. To determine the role of PAK in AD, we first quantified PAK by immunoblotting in homogenates from the parietal neocortex of subjects with a clinical diagnosis of no cognitive impairment (n = 12), mild cognitive impairment (n = 12), or AD (n = 12). A loss of total PAK, detected in the cortex of AD patients (-39% versus controls), was correlated with cognitive impairment (r(2) = 0.148, p = 0.027) and deposition of total and phosphorylated tau (r(2) = 0.235 and r(2) = 0.206, respectively), but not with Aβ42 (r(2) = 0.056). Accordingly, we found a decrease of total PAK in the cortex of 12- and 20-month-old 3xTg-AD mice, an animal model of AD-like Aβ and tau neuropathologies. To determine whether PAK dysfunction aggravates AD phenotype, 3xTg-AD mice were crossed with dominant-negative PAK mice. PAK inactivation led to obliteration of social recognition in old 3xTg-AD mice, which was associated with a decrease in cortical drebrin (-25%), but without enhancement of Aβ/tau pathology or any clear electrophysiological signature. Overall, our data suggest that PAK decrease is a consequence of AD neuropathology and that therapeutic activation of PAK may exert symptomatic benefits on high brain function.

The activity of ensembles of hippocampal place cells represents a hallmark of an animal’s spatial experience. The neuronal mechanisms that enable the rapid expression of novel place cell sequences are not entirely understood. Here we report that during sleep or rest, distinct sets of hippocampal temporal sequences in the rat preplay multiple corresponding novel spatial experiences with high specificity. These findings suggest that the place cell sequence of a novel spatial experience is determined, in part, by an online selection of a subset of cellular firing sequences from a larger repertoire of preexisting temporal firing sequences in the hippocampal cellular assembly network that become rapidly bound to the novel experience. We estimate that for the given context, the recorded hippocampal network activity has the capacity to preplay an extended repertoire of at least 15 future spatial experiences of similar distinctiveness and complexity.

The vitronectin receptor is a member of the integrin family of adhesion protein receptors and binds a broad spectrum of ligands, including fibronectin and fibrinogen in addition to vitronectin. We have generated four mAbs that recognize the murine alpha v beta 3 vitronectin receptor. Biochemical and expression analyses showed that two of the mAbs are specific for the alpha v chain, and two are specific for the beta 3 chain. The mAbs are effective blocking reagents and inhibited cell adhesion to vitronectin, fibrinogen, and fibronectin. Staining analysis revealed expression of alpha v and beta 3 on certain populations of murine thymocytes, splenocytes, and bone marrow cells. The expression of alpha v and beta 3 appeared to be modulated at specific stages of thymocyte development, suggesting a possible function for the alpha v beta 3 vitronectin receptor in T cell development.

To investigate the role of synaptic plasticity in the place-specific firing of the hippocampus, we have applied multiple electrode recording techniques to freely behaving mice with a CA1 pyramidal cell–specific knockout of the NMDAR1 gene. We have discovered that although the CA1 pyramidal cells of these mice retain place-related activity, there is a significant decrease in the spatial specificity of individual place fields. We have also found a striking deficit in the coordinated firing of pairs of neurons tuned to similar spatial locations. Pairs have uncorrelated firing even if their fields overlap. These results demonstrate that NMDA receptor–mediated synaptic plasticity is necessary for the proper representation of space in the CA1 region of the hippocampus.

The brain dopamine system is thought to be the major target for the neuropharmacological actions of psychomotor stimulants such as cocaine. To investigate the mechanisms of cocaine action, we used a genetic approach, the gene-targeting technique, and generated D1 dopamine receptor mutant mice. Locomotor activity analysis in response to cocaine indicates that, in contrast to control mice which showed a dose-dependent increase in locomotion, D1 receptor mutant mice exhibited a dose-dependent decrease, suggesting that D1 receptors play an essential role in mediating such effects. Extracellular single unit recording of dopamine sensitive nucleus accumbens neurons in the D1 receptor mutant mice and control mice revealed a marked reduction in the inhibitory effects of cocaine and dopamine on the generation of action potentials, suggesting that D1 receptors play a fundamental role in cocaine- and dopamine-mediated neurophysiological effects within the nucleus accumbens. From these analyses, we conclude that the D1 dopamine receptor plays essential roles in mediating these effects of cocaine. In the future, the use of this powerful genetic approach will be essential for elucidating the molecular components of the signal transduction pathway leading to anatomical, cellular and behavioral changes upon cocaine administration and dopamine neurotransmission.

Mucosal tissues of mice are enriched in T cells that express the gamma/delta T cell receptor. Since the function of these cells remains unclear, we have compared mucosal immune responses in gamma/delta T cell receptor-deficient (TCRdelta-/-) mice versus control mice of the same genetic background. The frequency of intestinal immunoglobulin (Ig) A plasma cells as well as IgA levels in serum, bile, saliva, and fecal samples were markedly reduced in TCRdelta-/- mice. The TCRdelta-/- mice produced much lower levels of IgA antibodies when immunized orally with a vaccine of tetanus toxoid plus cholera toxin as adjuvant. Conversely, the antigen-specific IgM and IgG antibody responses were comparable to orally immunized control mice. Direct assessment of the cells forming antibodies against the tetanus toxoid and cholera toxin antigens indicated that significantly lower numbers of IgA antibody-producing cells were present in the intestinal lamina propria and Peyer's patches of TCRdelta-/- mice compared with the orally immunized control mice. The selective reduction of IgA responses to ingested antigens in the absence of gamma/delta T cells suggests a specialized role for gamma/delta cells in mucosal immunity.

Since lumenal bacteria have been postulated to play an important role in the pathogenesis of inflammatory bowel disease (IBD), we investigated the humoral response to cecal aerobic bacterial antigens by Western blot analysis in TCR alpha -/- mice which spontaneously develop IBD. The sera from TCR alpha -/- mice revealed an alteration of the recognition pattern against aerobic bacterial antigens from polyclonal to oligoclonal with age. This alteration was not observed in TCR delta -/- and TCR alpha +/- mice. The alteration of the recognition pattern in TCR alpha -/- mice was associated with production of autoantibodies against tropomyosin and the development of IBD. The unique population of CD4+ TCR alpha -beta + cells in TCR alpha -/- mice may be involved in the recognition of these bacterial antigens and the absence of the alpha chain may result in the alteration of immune response.

Ca2+/phospholipid-dependent protein kinase has long been thought to play an important role in modulating synaptic efficacy. It has been shown previously that mice lacking the brain-specific gamma subtype of PKC display abnormal long-term potentiation (LTP), whereas ordinary synaptic transmission is unaffected. by the mutation. We now examine the effects of phorbol esters, which are nonselective activators of PKC, on synaptic modulation in these mutant mice. In wild-type mice, phorbol esters produce marked enhancement of synaptic transmission that is largely presynaptic in origin, an effect that has been thought to share mechanisms with LTP. In mutant mice, phorbol ester-mediated potentiation is normal despite the absence of the major PRC isoform. As in wild-type mice, this synaptic enhancement is at lease partly attributable to presynaptic changes. Our results demonstrate that the gamma isotype of PKC is not essential for phorbol ester-mediated synaptic facilitation, and place limitations on the possible roles of PKC in LTP.

We have generated mice deficient in HO-2, the major cerebral isoform of heme oxygenase, in order to assess the potential role of carbon monoxide as a retrograde messenger in hippocampal LTP. Cerebral HO catalytic activity was markedly reduced in the HO-2 mutant mice, yet no differences were found between wild types and mutants in gross neuroanatomical structure, in basal hippocampal synaptic transmission, or in the amount of potentiation produced by various LTP induction protocols. Furthermore, zinc protoporphyrin IX, an inhibitor of HO, had nearly identical inhibitory effects on LTP in wild-type and HO-2 mutant hippocampal slices. Our data indicate that carbon monoxide produced endogenously by HO is unlikely to be a neuromodulator required for LTP in the hippocampus.

The T-cell receptor (TCR) beta chain is instrumental in the progression of thymocyte differentiation from the CD4-CD8- to the CD4+CD8+ stage. This differentiation step may involve cell surface expression of novel CD3-TCR complexes. To facilitate biochemical characterization of these complexes, we established cell lines from thymic lymphomas originating from mice carrying a mutation in the p53 gene on the one hand and a mutation in TCR-alpha, TCR-beta, or the recombination activating gene 1 (RAG-1) on the other hand. The cell lines were CD4+CD8+ and appeared to be monoclonal. A cell line derived from a RAG-1 x p53 double mutant thymic lymphoma expressed low levels of CD3-epsilon, -gamma, and -delta on the surface. TCR-alpha x p53 double mutant cell lines were found to express complexes consisting of TCR-beta chains associated with CD3-epsilon, -gamma, and -delta chains and CD3-zeta zeta dimers. These lines will be useful tools to study the molecular structure and signal transducing properties of partial CD3-TCR complexes expressed on the surface of immature thymocytes.

Multifunctional Ca2+/calmodulin-dependent protein kinase II (CaMK) phosphorylates proteins pivotally involved in diverse neuronal processes and thereby coordinates cellular responses to external stimuli that regulate intracellular Ca2+ [Hanson, P. I. & Schulman, H. (1992) Annu. Rev. Biochem. 61, 559-664]. Despite extensive study, the impact of this enzyme on control of the excitability of neuron populations in the mammalian nervous system in situ is unknown. To address this question, we studied transgenic mice carrying a null mutation (-/-) for the alpha subunit of CaMK. In contrast to wild-type littermates, null mutants exhibit profound hyperexcitability, evident in epileptic seizures involving limbic structures including the hippocampus. No evidence of increased excitability was detected in mice carrying null mutations of the gamma isoform of protein kinase C, underscoring the specificity of the effect of CaMK. CaMK plays a powerful and previously underappreciated role in control of neuronal excitability in the mammalian nervous system. These insights have important implications for analyses of mechanisms of epilepsy and, perhaps, learning and memory.

Although T cells bearing gamma delta T-cell receptors have long been known to be present in the epithelial lining of many organs, their specificity and function remain elusive. In the present study, we examined the intestinal epithelia of T-cell-receptor mutant mice, which were deficient in either gamma delta T cells or alpha beta T cells, and of normal littermates. The absence of gamma delta T cells was associated with a reduction in epithelial cell turnover and a downregulation of the expression of major histocompatibility complex class II molecules. No such effects were observed in alpha beta T-cell-deficient mice. These findings indicate that intraepithelial gamma delta T cells regulate the generation and differentiation of intestinal epithelial cells.

Calcium/phospholipid-dependent protein kinase (protein kinase C, PKC) has been suggested to play a role in the sensitivity of gamma-aminobutyrate type A (GABAA) receptors to ethanol. We tested a line of null mutant mice that lacks the gamma isoform of PKC (PKC gamma) to determine the role of this brain-specific isoenzyme in ethanol sensitivity. We found that the mutation reduced the amount of PKC gamma immunoreactivity in cerebellum to undetectable levels without altering the levels of the alpha, beta I, or beta II isoforms of PKC. The mutant mice display reduced sensitivity to the effects of ethanol on loss of righting reflex and hypothermia but show normal responses to flunitrazepam or pentobarbital. Likewise, GABAA receptor function of isolated brain membranes showed that the mutation abolished the action of ethanol but did not alter actions of flunitrazepam or pentobarbital. These studies show the unique interactions of ethanol with GABAA receptors and suggest protein kinase isoenzymes as possible determinants of genetic differences in response to ethanol.

Correction to: Neuropsychopharmacology Reviews (2016) 41, 370–371; doi:10.1038/npp.2015.264 In this paper, the first reference in the list should read: Liu X, Ramirez S, Pang PT, Puryear CB, Govindarajan A, Deisseroth K et al (2012). Optogenetic stimulation of a hippocampal engram activates fear memory recall.

Mice lacking T cells with alpha beta TCR (TCR beta-/-) or gamma delta TCR (TCR delta-/-) were infected with the erythrocytic stages of the malaria parasite, Plasmodium chabaudi chabaudi (AS). Mice without gamma delta T cells could control and reduce a primary infection of P. chabaudi with a slight delay in the time of clearance of the acute phase of infection and significantly higher recrudescent parasitaemias compared with control intact mice. TCR delta -/- mice had higher levels of both serum Ig and malaria-specific antibodies of the isotypes IgG3 and IgG1 compared with control mice. TCR beta -/- mice, despite a striking increase in NK1.1+ cells and the presence of gamma delta T cells, were unable to clear their infection. Although the plasma of TCR beta -/- mice contained all Ig isotypes before and during a primary infection, they were unable to produce significant levels of malaria-specific IgG antibodies, suggesting that in the absence of alpha beta T cells gamma delta T cells are not able to provide efficient help for antibody production.

Susumu Tonegawa is a Japanese molecular biologist who won the Nobel Prize for Physiology or Medicine in 1987 for his discovery of "the genetic principle for generation of antibody diversity". Susumu Tonegawa is also famous for his important contributions on neuroscience, who explored the molecular and cellular mechanisms underlying learning and memory. In this article, the life and research on Susumu Tonegawa was introduced.

In the developing nervous system synaptic refinement, typified by the neuromuscular junction where supernumerary connections are eliminated by axon retraction leaving the postsynaptic target innervated by a single dominant input, critically regulates neuronal circuit formation. Whether such competition-based pruning continues in established circuits of mature animals remains unknown. This question is particularly relevant in the context of adult neurogenesis where newborn cells must integrate into preexisting circuits, and thus, potentially compete with functionally mature synapses to gain access to their postsynaptic targets. The hippocampus plays an important role in memory formation/retrieval and the dentate gyrus (DG) subfield exhibits continued neurogenesis into adulthood. Therefore, this region contains both mature granule cells (old GCs) and immature recently born GCs that are generated throughout adult life (young GCs), providing a neurogenic niche model to examine the role of competition in synaptic refinement. Recent work from an independent group in developing animals indicated that embryonically/early postnatal generated GCs placed at a competitive disadvantage by selective expression of tetanus toxin (TeTX) to prevent synaptic release rapidly retracted their axons, and that this retraction was driven by competition from newborn GCs lacking TeTX. In contrast, following 3–6 months of selective TeTX expression in old GCs of adult mice we did not observe any evidence of axon retraction. Indeed ultrastructural analyses indicated that the terminals of silenced GCs even maintained synaptic contact with their postsynaptic targets. Furthermore, we did not detect any significant differences in the electrophysiological properties between old GCs in control and TeTX conditions. Thus, our data demonstrate a remarkable stability in the face of a relatively prolonged period of altered synaptic competition between two populations of neurons within the adult brain.

Uterine leiomyosarcoma (ULMS) develops more often in the muscle tissue layer of the uterine body than in the uterine cervix. The development of gynecologic tumors is often correlated with female hormone secretion; however, the development of uterine ULMS is not substantially correlated with hormonal conditions, and the risk factors are not yet known. Importantly, a diagnostic-biomarker which distinguishes malignant ULMS from benign tumor leiomyoma (LMA) is yet to be established. Accordingly, it is necessary to analyze risk factors associated with uterine ULMS, to establish a treatment method. Proteasome low-molecular mass polypeptide 2(LMP2)/b1i-deficient mice spontaneously develop uterine LMS, with a disease prevalence of ~40% by 14 months of age. We found LMP2/b1i expression to be absent in human LMS, but present in human LMA. Therefore, defective-LMP2/b1i expression may be one of the risk factors for ULMS. LMP2/b1i is a potential diagnostic-biomarker for uterine ULMS, and may be a targeted-molecule for a new therapeutic approach.

Adult-born granule cells (GCs), a minor population of cells in the hippocampal dentate gyrus, are highly active during the first few weeks following functional integration into the neuronal network (young GCs), distinguishing them from less active older adult-born GCs and the major population of dentate GCs generated developmentally (together, old GCs). We created a transgenic mouse in which output of old GCs was specifically inhibited while leaving a substantial portion of young GCs intact. These mice exhibited enhanced or normal pattern separation between similar contexts that was reduced following removal of young GCs by X-ray irradiation. Furthermore, mutant mice exhibited deficits in rapid pattern completion. Therefore, pattern separation of similar contexts requires adult-born young GCs while old GCs are unnecessary, whereas older GCs contribute to the rapid recall by pattern completion. Our data suggest that as adult-born GCs age, their function switches from pattern separation to rapid pattern completion.

The brain dopaminergic system is a critical modulator of basal ganglia function and plasticity. To investigate the contribution of the dopamine D1 receptor to this modulation, we have used gene targeting technology to generate D1 receptor mutant mice. Histological analyses suggested that there are no major changes in general anatomy of the mutant mouse brains, but indicated that the expression of dynorphin is greatly reduced in the striatum and related regions of the basal ganglia. The mutant mice do not respond to the stimulant and suppressive effects of D1 receptor agonists and antagonists, respectively, and they exhibit locomotor hyperactivity. These results suggest that the D1 receptor regulates the neurochemical architecture of the striatum and is critical for the normal expression of motor activity.

mGluR1 mutant mice are viable but show characteristic cerebellar symptoms such as ataxic gait and intention tremor. The anatomy of the cerebellum is not overtly disturbed. Excitatory synaptic transmission from parallel fibers (PFs) to Purkinje cells and that from climbing fibers (CFs) to Purkinje cells appear to be functional, and voltage-gated Ca2+ channels of Purkinje cells are normal. Both PF and CF synapses display normal short-term synaptic plasticity to paired stimuli. By marked contrast, long-term depression (LTD) is clearly deficient and conditioned eyeblink response is impaired. We conclude that mGluR1 is required for the induction of LTD and that the ataxic behavior and impaired eyeblink conditioning of the mGluR1 mutant mice are primarily due to deficient LTD.

Background Calcium influx into postsynaptic dendritic spines can, depending on circumstances, activate three forms of synaptic plasticity: long-term potentiation (LTP), short-term potentiation (STP) and long-term depression (LTD). The increased postsynaptic calcium concentrations that trigger all three forms of plasticity should activate the α isoform of calcium–calmodulin kinase type II ( α CaMK II), which is present at high levels just below the postsynaptic membrane. Earlier experiments have implicated α CaMK II in the regulation or induction of LTP, but no information is available on the possible role of this enzyme in the two other forms of synaptic plasticity, STP and LTD. Results We used mice that lack the gene for α CaMK II to investigate the role of this enzyme in synaptic plasticity. Field potential recordings from hippocampal slices taken from mutant mice show that STP and LTD are, like LTP, absent or markedly attenuated in the absence of α CaMK II. A brief form of synaptic modification — post-tetanic potentiation (PTP) — is, however, intact in the absence of this enzyme. Conclusion It appears likely that α CaMK II is involved in the production or global regulation of all three forms of synaptic plasticity. We propose that the activation of this enzyme is a common step in the induction of LTP and STP, and that α CaMK II activity is required for the normal production of LTD.

The processes of positive and negative selection during thymic development shape the repertoires of antigen specificities displayed by T cells. This rids the animal of potentially autoreactive T cells and, at the same time, ensures that they are capable of major histocompatibility complex (MHC)-restricted recognition of antigen. Paradoxically, both processes involve the engagement of the T-cell recepetor (TCR) on immature thymocytes with peptide/MHC complexes expressed on thymic stromal cells. Here, Philip Ashton-Rickardt and Susumu Tonegawa suggest that the critical parameter determining the outcome of this interaction is the number of TCRs occupied by peptide/MHC complexes and that this, in turn, is determined by the avidity of the TCR-MHC interaction: low avidity resulting in positive selection and high avidity resulting in negative selection.

We have analyzed the specificity and function of natural killer (NK) cells in mice with a homozygous deletion of the major histocompatibility complex (MHC)-encoded transporter gene associated with MHC class I-restricted antigen presentation (Tap-1). These mice express very low levels of class I molecules at the cell surface, and these molecules are either devoid of peptide or occupied only by TAP-independent peptides. NK cells in Tap-1 -/- mice, through normal in number, appeared tolerant toward autologous Tap-1 -/- Con A-activated blasts, Tap-1 -/- as well as allogeneic BALB/c bone marrow cells, and RMA-S tumor cell grafts. In contrast, they killed YAC-1 cells as efficiently as did NK cells from wild-type mice. Defective Tap-1 expression was sufficient to render nontransformed target cells sensitive to NK cell-mediated lysis. It is concluded that proper expression of TAP molecules is necessary for normal development of NK cells, as well as for rendering target cells resistant to NK cell-mediated lysis. These results support the hypothesis that class I molecules of the MHC influence the sensitivity of target cells to lysis by NK cells, as well as the development of the NK cell repertoire.

Mice with a homozygous deletion in their Tap-1 gene (-/- mice) express very low levels of cell membrane major histocompatibility complex class I molecules and have < 1% peripheral CD8+ T cells. We show that these -/- mice but not their +/- littermates display strong primary syngeneic anti-H-2Kb and -Db-specific responses mediated by CD8+ T cells. These responses are augmented by in vivo priming. Further, -/- mice primed in vivo with H-2d alloantigens generate an anti-H-2d response which appears nearly as strong as that found in +/- littermates. Both -/- anti-H-2b and anti-H-2d T cells do not recognize target cells from Tap-1 -/- animals or Tap-2-deficient RMA-S cells. Thus, some CD8+ anti-self and alloreactive T cells can be selected in the absence of Tap proteins.

Positive and negative selection of a lymphocytic choriomeningitis virus (LCMV) peptide-specific, H-2Db-restricted T cell clone (P14) was studied using TAP1- and TAP1+ mice transgenic for P14 T cell receptor (TCR) alpha and beta genes. Positive selection of transgenic CD8+ P14 cells was impaired in TAP1- mice. Addition of the LCMV peptide to TAP1- fetal thymic organ cultures (FTOCs) at low and high concentrations induced positive and negative selection of CD8+ P14 cells, respectively, while addition of the same peptide to TAP1+ FTOCs induced negative selection even at low concentrations. Both types of selection were peptide specific. Thus, a critical parameter that controls the fate of a thymocyte seems to be the number of TCRs engaged with complexes of peptide and major histocompatibility complex. When this number is low, positive selection occurs, and when it is high, negative selection takes place. These findings support a differential avidity model of T cell selection.

Associating temporally discontinuous elements is crucial for the formation of episodic and working memories that depend on the hippocampal-entorhinal network. However, the neural circuits subserving these associations have remained unknown. The layer III inputs of the entorhinal cortex to the hippocampus may contribute to this process. To test this hypothesis, we generated a transgenic mouse in which these inputs are specifically inhibited. The mutant mice displayed significant impairments in spatial working-memory tasks and in the encoding phase of trace fear-conditioning. These results indicate a critical role of the entorhinal cortex layer III inputs to the hippocampus in temporal association memory.

The majority of smooth muscle tumors found in the uterus are benign, but uterine leiomyosarcomas (LMSs) are extremely malignant, with high rates of recurrence and metastasis. The development of gynecologic tumors is often correlated with female hormone secretion; however, the development of uterine LMS is not substantially correlated with hormonal conditions, and the risk factors are not clearly understood. The presentation of antigenic peptides by major histocompatibility complex (MHC) class I molecules is important for tumor rejection by cytotoxic T-lymphocytes (CTLs). Such antigenic peptides are generated as a result of the degradation of intracellular proteins by the proteasome pathway, a process that is influenced by the interferon (IFN)-γ-inducible low molecular mass polypeptide-2 (LMP2) subunit of the 20S proteasome. Homozygous deficient mice for LMP2 are now known to spontaneously develop uterine LMS. LMP2 expression is reportedly absent in human uterine LMS, but present in human myometrium. Further studies revealed a few infiltrating CD56+ NK cells in human uterine LMS tissues. This review aims at summarizing recent insights into the regulation of NK cell function and the T cell-mediated immune system as tumor immune surveillance, first attempts to exploit NK cell activation to improve immunity to tumors.