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A significant fraction of the total calcium/calmodulin-dependent protein kinase II (CaMKII) activity in neurons is associated with synaptic connections and is present in nerve terminals, thus suggesting a role for CaMKII in neurotransmitter release. To determine whether CaMKII regulates neurotransmitter release, we generated and analyzed knockout mice in which the dominant [alpha]-isoform of CaMKII was specifically deleted from the presynaptic side of the CA3-CA1 hippocampal synapse. Conditional CA3 [alpha]-CaMKII knockout mice exhibited an unchanged basal probability of neurotransmitter release at CA3-CA1 synapses but showed a significant enhancement in the activity-dependent increase in probability of release during repetitive presynaptic stimulation, as was shown with the analysis of unitary synaptic currents. These data indicate that [alpha]-CaMKII serves as a negative activity-dependent regulator of neurotransmitter release at hippocampal synapses and maintains synapses in an optimal range of release probabilities necessary for normal synaptic operation.

Results point out that the calcium-dependent protein phosphatase, calcineurin, of the mouse forebrain mediates the bidirectional synaptic plasticity and differentiates between working/episodic-like memory and reference memory. Mutant mice with disrupted calcineurin show impairment in hippocampus-dependent working and episodic-like memory tasks.

Research has been conducted on the calcium/calmodulin-dependent protein kinase IV. The role of this kinase in the neuronal plasticity and memory has been investigated via the generated transgenic mice with the dominant-negative calcium/calmodulin-dependent protein kinase IV form expression restricted to the postnatal forebrain and the results are presented.

Brain-derived neurotrophic factor (BDNF), which regulates maturation of inhibition and the critical period of plasticity in the visual cortex of mice, is discussed. Visual cortex maturation is influenced by visual experience during an early postnatal period. In transgenic mice in which the postnatal increase of BDNF was speeded up, this was studied. Maturation of GABAergic innervation and inhibition was speeded up and the age-dependent decline of cortical long-term potentiation induced by white matter stimulation happened sooner. Precocious development of visual acuity and an earlier end for the critical period for ocular dominance plasticity was seen.

In this study, we compared the influence of two peptides on the selection of CD8[Alpha][Alpha] and CD8[Alpha][Beta] intra-epithelial 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-2[D.sup.b]. 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 CD[3.sup.+]CD[8.sup.+] 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.sup.+][CD8.sup.+] thymocytes was not rescued by these transgenic peptides. However, the high-affinity NP68 peptide induced maturation of CD8[Alpha][Alpha] IEL, whereas the low-affinity NP34 peptide stimulated development of both CD8[Alpha][Beta] and CD8[Alpha][Alpha] IEL, but in smaller numbers. When both peptides were present, CD8[Alpha][Beta] T cells failed to develop and the number of CD8[Alpha][Alpha] 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.

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[prime],5[prime] monophosphate elevations evoked by electrical field stimulation were diminished markedly in both nNO[S.sup.[Delta]/[Delta]] and H[O-2.sup.[Delta]/[Delta]] mice. In wild-type mice, NOS inhibitors and HO inhibitors partially inhibited nonadrenergic noncholinergic relaxation. In nNO[S.sup.[Delta]/[Delta]] animals, NOS inhibitors selectively lost their efficacy, and HO inhibitors were inactive in H[O-2.sup.[Delta]/[Delta]] animals.

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 (PKC[Gamma]) 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, PKC[Gamma] 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., Neuropathic pain is a devastating consequence of nerve injury that is characterized by spontaneous, often burning, pain, an exaggerated response to painful stimuli (hyperalgesia), and pain in response to normally [...]

Most of the [Gamma][Delta] T cells in the intestinal epithelium of normal mice use the V[Gamma]1 or the V[Gamma]7 gene segments. However, the relative proportions of [Gamma][Delta] intraepithelial lymphocytes expressing either the V[Gamma]I or the V[Gamma]7 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 [Gamma][Delta] T cell subsets in the intestinal epithelium, we used available anti-T cell antigen receptor (TCR) V region-specific mAbs against V[Gamma]l, V[Gamma]4, V[Gamma]7, and V[Delta]4 to examine the TCR repertoire of intraepithelial [Gamma][Delta] lymphocytes in a set of (C57BL/6 x DBA/2) recombinant inbred strains. Our results show that the representation of different V[Gamma] and V[Delta] gene products among [Gamma][Delta] intestinal intraepithelial lymphocytes is under a complex genetic control with a marked influence by genes closely linked to the TCR[Gamma], TCR[Delta], and major histocompatibility complex loci.

The normal physiological role of the Presenilin-1 (PS1) was analyzed in PS1 homozygous mutant mice. Homozygous mutations in the PS1 gene in mice that were derived from four independent embryonic stem cell lines induced death after natural birth or caesarean-section. The PS1 homozygous mutant mice also exhibited gross skeletal and central nervous system abnormalities . Furthermore, brain abnormalities in the mutants was caused by reduced number of progenitor cells and neurons in the ventrolateral region of the ventricular zone.

The use of genetically engineered mice has been a major development in neuroscience research. Genetic engineering is an undoubtedly powerful technique; however, the value of this approach has been debated, particularly in relation to its use to probe the underlying bases of complex behaviors, such as memory. A recent new development of the technique is the ability to target a specific gene knockout to a particular subregion or even to specific and limited cell types of the mouse brain. An example of this approach is the knockout of the NMDARI gene in only CAI-pyramidal cells of the hippocampus. The resulting animals can be tested by several methods, including in vivo multielectrode recording during behavioral tasks. The data provide strong evidence in favor of the notion that NMDA receptor-dependent synaptic plasticity at CAI synapses is required for both the acquisition of spatial memory and the formation of normal CAI place fields. This relationship suggests that robust place fields may be essential for spatial memory.

The neurons of the central nervous system of mammals usually lose their regenerative ability after a certain point in their development. Unlike retinal ganglion cells (RGCs) of embryos, RGCs from older animals cannot grow axons into tectum of any age. The role of the proto-oncogene bcl-2 in this developmental shift is examined. It is found to stimulate the growth and regenerate retinal axons, although this effect does not appear to be indirectly related to the proto-oncogene's anti-apoptotic activity.

A cell-type restricted gene knockout technology for mice was developed based on the phage P1-derived Cre/loxP recombination-based gene deletion system. The forebrain restricted gene knockout technology involves the production of Cre transgenic mice by Cre/loxP recombination is dependent on the nature of the transcription promoter that regulates Cre gene expression. The floxed stop and lacZ sequences were also utilized to determine the subregional specificity of Cre/lox recombination.

A novel method based on the Cre/loc recombination system from phage P1 was utilized for the restricted deletion of the (N-methyl-D-aspartate receptor) NMDAR1 gene in the pyramidal cells of the hippocampal CA1 region of mice. The CA1-restricted NMDAR1 mutants developed normal brains but the slow component of the excitatory postsynaptic current was absent in their CA1-knock out cells. Furthermore, the mutant mice exhibited deficient learning abilities and spatial memory indicating the role of NMDA in the formation of specific forms of memory.

The hippocampal activity of mutant mice exhibiting CA1 restricted deletion of the N-methyl-D-aspartate receptor subunit 1 (NMDAR1) gene was analyzed by utilizing multielectrode recording techniques. The CA1 restricted NMDAR1 mutants exhibited normal neuronal firing activity characterized by rhythmic modulations of the firing rate with a theta frequency of 8-10 Hertz. However, the NMDAR1 mutants exhibited deficient neuronal firing during the performance of spatial tasks indicating the role of NMDAR1 in spatial memory and synaptic plasticity.

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.

Mutant mice deficient in protein kinase C-gamma are unable to coordinate motor activity but learn discrete motion elements such as eyeblinking conditioning. Cerebellar long-term depression is unaffected in the mutants.The motor coordination defects are probably due to the presence of excess multiple climbing fibers in the Purkinje cells of the mutants.

Protein kinase C-gamma (PKC-gamma) may function as a downstream element in eliminating excess climbing fiber (CF) synapses. Adult mutant mice lacking PKC-gamma show multiple CFs in 41% of the Purkinje cells (PCs) unlike the wild-type mice whose PCs have single CF innervation. The mutants have a normal anatomy and normal transmission of excitatory stimuli in CF-PC and the parallel fiber-PC synapses. The mutants show normal regression of multiple CF innervation until two weeks after birth followed by a reduction in regression.

Multifunctional Ca2+/ calmodulin-dependant protein kinase II (CaMK) appears to play a large role in the control of neuron excitability in mammalian nervous systems, with a possible role in epilepsy. Studies on the affects of CaMK alpha-subunit null mutations in transgenic mice found that such mutations led to high excitability and and epileptic seizures. Similar mutation of the gamma-subunit had no affects on excitability.

The intraepithelial gamma-delta T cells may regulate the generation and differentiation of intestinal epithelial cells. Researchers examined the intestinal epithelium of T-cell-mutant mice lacking either gamma-delta T cells or alpha-beta T cells. The tissue of mice without gamma-delta T cells exhibited less cell turnover a lower regulatory expression of major histocompatibility complex class II molecules. Alpha-beta mouse epithelium did not show these characteristics.

Calcium/phospholipid-dependent protein kinase (protein kinase C, PKC), a brain-related isoenzyme, has a major role in the interaction of gamma-aminobutyrate type A (GABA sub A) receptors with ethanol. Accordingly, a study with null mutant mice lacking the gamma isoform of PKC shows the decreased action of ethanol on loss of righting reflex and hypothermia but the mice display normal responses to drugs such as flunitrazepam and pentobarbital.

A study of the involvement of the dopamine D1 receptor in cocaine-induced hyperactivity using gene targeting-produced dopamine receptor mutant mice reveals a decrease in locomotion in the mutant mice that is dose-dependent. Electrophysiological studies reveal the complete elimination of the cocaine-induced inhibition of the production of action potentials by the dopamine-sensitive nucleus accumbens neurons. The mutant mice do not possess dopamine and D1 and D2 agonist inhibitory effects but retain the inhibitory effects of serotonin. The mice also exhibit normal D2-like dopamine receptor binding.

Histological studies to determine the function of brain dopamine D1 receptor in D1 receptor mutant mice reveals that the D1 receptor is vital for normal motor activity levels and modulates the neurochemical structure of the striatum. The mutant mice display a decrease in dymorphin expression in the basal ganglia and striatum but do not display significant changes in the anatomy of the brain. D1 receptor antagonists and agonists fail to trigger the mutant mice that also display locomotor hyperactivity.

mGLuR1-absent mice exhibit ataxic cerebellar symptoms and defective long term depression (LTD) and conditioned eyeblink response. They exhibit normal cerebellar anatomy, functional excitatory synaptic transmission from parallel and climbing fibers to the Purkinje cells and normal voltage-gated Ca+ channels of Purkinje cells. Observations on the mutant mice suggest that LTD activation requires mGLuR1 and that LTD deficiency leads to defective eyeblinking and ataxic symptoms.

The generation of a new strain of mutant mice with a deletion in the gene that encodes metabotropic glutamate receptor 1 enables the study of its involvement in learning and memory through its role in synaptic plasticity and transmission. The mutant exhibits deficiency in context-specific associative learning and diminished long term potentiation, despite normal levels of short term potentiation, normal hippocampal anatomy and synaptic transmission.

Mice deficient for the gene encoding α-calcium-calmodulin-dependent kinase II (α-CaMKII knockout mice) provide a promising tool to link behavioral and cellular abnormalities with a specific molecular lesion. The heterozygous mouse exhibited a well-circumscribed syndrome of behavioral abnormalities, consisting primarily of a decreased fear response and an increase in defensive aggression, in the absence of any measured cognitive deficits. Unlike the heterozygote, the homozygote displayed abnormal behavior in all paradigms tested. At the cellular level, both extracellular and whole-cell patch clamp recordings indicated that serotonin release in putative serotonergic neurons of the dorsal raphe was reduced. Thus, α-CaMKII knockout mice, in particular the heterozygote, may provide a model for studying the molecular and cellular basis underlying emotional disorders involving fear and aggression., The recently developed mouse gene knockout technology has allowed us to use a multidisciplinary approach to analyze neurobiological abnormalities at the behavioral and cellular levels. Here we tested mutant mice, [...]

Activated myelin basic protein (MBP)-specific T lymphocytes occur in the brain of T cell receptor (TCR) non-transgenic (T/R-) and transgenic (T/R+) mice suffering from autoimmune encephalomyelitis (EAE). In the absence of other lymphocytes, MBP-specific CD4 T cells regulate EAE development. Nontransgenic lymphocytes offer protection against spontaneous EAE, although both T/R- and T/R+ mice have significantly encephalitogenic CD4 T cells. There is 100% susceptibility to EAE in mice expressing transgenic TCR.

Study of allogeneic and anti-self class I repertoire in Tap-1 mice reveals the possibility of selecting positive self- and alloreactive CD8+ T cells when Tap proteins are non-existent. The mice demonstrated syngeneic anti-H-2Kb and -Db-specific responses in the presence of CD8+ T cells. CD8+ T cell repertoire is sensitive to positive and negative selection.

The specificity of TCR-MHC interaction during positive selection is decided by the recognition of peptides by TCRs. The number of TCRs involved in peptide-MHC complexes is decided by the avidity between the two cell surface components, which in turn decides the fate of thymocytes. Moderate number of TCR leads to positive selection, while a high number results in negative selection.

Genomic Southern blot analysis of muted mice resulting from gene targeting of NMDAR1 subunit of N-methyl-D-aspartate (NMDA) revealed that while targeting and topographic projection of trigeminal afferents and postsynaptic neuron response to stimulation were normal, whisker-specific patterns did not appear due to the absence of NMDA receptors. The NMDA receptor influenced periphenyl-related neural patterns in mammals.

Gamma delta T cells confer immunity against liver and blood malarial stage to alpha beta T cell-deficient mice. Murine gamma delta T cell mutants that act against microbes define T cell recognition of the antigens. The incorporation of germ-line mutations in either the TCR alpha beta or gamma delta chain mice genes yields the alpha beta T cell lacking test mice.

PKC-gamma mutant mice, characterized by modified long-term potentiation (LTP) of synaptic transmission on the hippocampus, exhibit corresponding deficits in spatial and contextual learning behavior. Disruption of long-term depression may also cause deficits in learning behavior. Presence of hippocampal CA1 LTP, resulting from tetanic stimulator, is not a prerequisite for spatial and contextual learning.