Restoration of synaptic plasticity and learning in young and aged NCAM-deficient mice by enhancing neurotransmission mediated by GluN2A-containing NMDA receptors.

Neural cell adhesion molecule (NCAM) is the predominant carrier of the unusual glycan polysialic acid (PSA). Deficits in PSA and/or NCAM expression cause impairments in hippocampal long-term potentiation and depression (LTP and LTD) and are associated with schizophrenia and aging. In this study, we show that impaired LTP in adult NCAM-deficient (NCAM(-/-)) mice is restored by increasing the activity of the NMDA subtype of glutamate receptor (GluN) through either reducing the extracellular Mg2+ concentration or applying d-cycloserine (DCS), a partial agonist of the GluN glycine binding site. Pharmacological inhibition of the GluN2A subtype reduced LTP to the same level in NCAM(-/-) and wild-type (NCAM(+/+)) littermate mice and abolished the rescue by DCS in NCAM(-/-) mice, suggesting that the effects of DCS are mainly mediated by GluN2A. The insufficient contribution of GluN to LTD in NCAM(-/-) mice was also compensated for by DCS. Furthermore, impaired contextual and cued fear conditioning levels were restored in NCAM(-/-) mice by administration of DCS before conditioning. In 12-month-old NCAM(-/-), but not NCAM(+/+) mice, there was a decline in LTP compared with 3-month-old mice that could be rescued by DCS. In 24-month-old mice of both genotypes, there was a reduction in LTP that could be fully restored by DCS in NCAM(+/+) mice but only partially restored in NCAM(-/-) mice. Thus, several deficiencies of NCAM(-/-) mice can be ameliorated by enhancing GluN2A-mediated neurotransmission with DCS.

Experimental febrile seizures are precipitated by a hyperthermia-induced respiratory alkalosis.

Febrile seizures are frequent during early childhood, and prolonged (complex) febrile seizures are associated with an increased susceptibility to temporal lobe epilepsy. The pathophysiological consequences of febrile seizures have been extensively studied in rat pups exposed to hyperthermia. The mechanisms that trigger these seizures are unknown, however. A rise in brain pH is known to enhance neuronal excitability. Here we show that hyperthermia causes respiratory alkalosis in the immature brain, with a threshold of 0.2-0.3 pH units for seizure induction. Suppressing alkalosis with 5% ambient CO2 abolished seizures within 20 s. CO2 also prevented two long-term effects of hyperthermic seizures in the hippocampus: the upregulation of the I(h) current and the upregulation of CB1 receptor expression. The effects of hyperthermia were closely mimicked by intraperitoneal injection of bicarbonate. Our work indicates a mechanism for triggering hyperthermic seizures and suggests new strategies in the research and therapy of fever-related epileptic syndromes.

GluK1 inhibits calcium dependent and independent transmitter release at associational/commissural synapses in area CA3 of the hippocampus.

CA3 pyramidal cells receive three main excitatory inputs: the first one is the mossy fiber input, synapsing mainly on the proximal apical dendrites. Second, entorhinal cortex cells form excitatory connections with CA3 pyramidal cells via the perforant path in the stratum lacunosum moleculare. The third input involves the ipsi-and contralateral connections, termed the associational/commissural (A/C) pathway terminating in the stratum radiatum of CA3, thus forming a feedback loop within this region. Since this excitatory recurrent synapse makes the CA3 region extremely prone to seizure development, understanding the regulation of synaptic strength of this connection is of crucial interest. Several studies suggest that kainate receptors (KAR) play a role in the regulation of synaptic strength. Our aim was to characterize the influence of KAR on A/C synaptic transmission: application of ATPA, a selective agonist of the GluK1 KAR, depressed the amplitude fEPSP without affecting the size of the fiber volley. Blockade of GABA receptors had no influence on this effect, arguing against the influence of interneuronal KARs. Pharmacological and genetic deletion studies could show that this effect was selectively due to GluK1 receptor activation. Several lines of evidence, such as PPF changes, coefficient of variance-analysis and glutamate uncaging experiments strongly argue for a presynaptic locus of suppression. This is accompanied by an ATPA-mediated reduction in Ca(2+) influx at excitatory synaptic terminals, which is most likely mediated by a G-Protein dependent mechanism, as suggested by application of pertussis toxin. Finally, analysis of miniature EPSCs in the presence and absence of extracellular Ca(2+) suggest that presynaptic KAR can also reduce transmitter release downstream and therefore independent of Ca(2+) influx.

Arc/Arg3.1 is essential for the consolidation of synaptic plasticity and memories.

Arc/Arg3.1 is robustly induced by plasticity-producing stimulation and specifically targeted to stimulated synaptic areas. To investigate the role of Arc/Arg3.1 in synaptic plasticity and learning and memory, we generated Arc/Arg3.1 knockout mice. These animals fail to form long-lasting memories for implicit and explicit learning tasks, despite intact short-term memory. Moreover, they exhibit a biphasic alteration of hippocampal long-term potentiation in the dentate gyrus and area CA1 with an enhanced early and absent late phase. In addition, long-term depression is significantly impaired. Together, these results demonstrate a critical role for Arc/Arg3.1 in the consolidation of enduring synaptic plasticity and memory storage.

Differential involvement of the extracellular 6-O-endosulfatases Sulf1 and Sulf2 in brain development and neuronal and behavioural plasticity.

The extracellular sulfatases Sulf1 and Sulf2 remove specific 6-O-sulfate groups from heparan sulfate, thereby modulating numerous signalling pathways underlying development and homeostasis. In vitro data have suggested that the two enzymes show functional redundancy. To elucidate their in vivo functions and to further address the question of a putative redundancy, we have generated Sulf1- and Sulf2-deficient mice. Phenotypic analysis of these animals revealed higher embryonic lethality of Sulf2 knockout mice, which can be associated with neuroanatomical malformations during embryogenesis. Sulf1 seems not to be essential for developmental or postnatal viability, as mice deficient in this sulfatase show no overt phenotype. However, neurite outgrowth deficits were observed in hippocampal and cerebellar neurons of both mutant mouse lines, suggesting that not only Sulf2 but also Sulf1 function plays a role in the developing nervous system. Behavioural analysis revealed differential deficits with regard to cage activity and spatial learning for Sulf1- and Sulf2-deficient mouse lines. In addition, Sulf1-specific deficits were shown for synaptic plasticity in the CA1 region of the hippocampus, associated with a reduced spine density. These results reveal that Sulf1 and Sulf2 fulfil non-redundant functions in vivo in the development and maintenance of the murine nervous system.

Conditional ablation of the neural cell adhesion molecule reduces precision of spatial learning, long-term potentiation, and depression in the CA1 subfield of mouse hippocampus.

NCAM, a neural cell adhesion molecule of the immunoglobulin superfamily, is involved in neuronal migration and differentiation, axon outgrowth and fasciculation, and synaptic plasticity. To dissociate the functional roles of NCAM in the adult brain from developmental abnormalities, we generated a mutant in which the NCAM gene is inactivated by cre-recombinase under the control of the calcium-calmodulin-dependent kinase II promoter, resulting in reduction of NCAM expression predominantly in the hippocampus. This mutant (NCAMff+) did not show the overt morphological and behavioral abnormalities previously observed in constitutive NCAM-deficient (NCAM-/-) mice. However, similar to the NCAM-/- mouse, a reduction in long-term potentiation (LTP) in the CA1 region of the hippocampus was revealed. Long-term depression was also abolished in NCAMff+ mice. The deficit in LTP could be rescued by elevation of extracellular Ca2+ concentrations from 1.5 or 2.0 to 2.5 mm, suggesting an involvement of NCAM in regulation of Ca2+-dependent signaling during LTP. Contrary to the NCAM-/- mouse, LTP in the CA3 region was normal, consistent with normal mossy fiber lamination in NCAMff+ as opposed to abnormal lamination in NCAM-/- mice. NCAMff+ mutants did not show general deficits in short- and long-term memory in global landmark navigation in the water maze but were delayed in the acquisition of precise spatial orientation, a deficit that could be overcome by training. Thus, mice conditionally deficient in hippocampal NCAM expression in the adult share certain abnormalities characteristic of NCAM-/- mice, highlighting the role of NCAM in the regulation of synaptic plasticity in the CA1 region.

Impairment of L-type Ca2+ channel-dependent forms of hippocampal synaptic plasticity in mice deficient in the extracellular matrix glycoprotein tenascin-C.

The extracellular matrix glycoprotein tenascin-C (TN-C) has been suggested to play important functional roles during neural development, axonal regeneration, and synaptic plasticity. We generated a constitutively TN-C-deficient mouse mutant from embryonic stem cells with a floxed tn-C allele, representing a standard for future analysis of conditionally targeted mice. The gross morphology of the CNS was not detectably affected, including no evidence for perturbed nerve cell migration, abnormal oligodendrocyte distribution, or defective myelination. Despite the apparent normal histology of the hippocampus and normal performance in the water maze, theta-burst stimulation (TBS) of Schaffer collaterals elicited reduced long-term potentiation (LTP) in the CA1 region of TN-C-deficient mutants, as compared with wild-type littermates. However, high-frequency stimulation evoked normal LTP not only in CA1, but also at mossy fiber-CA3 and medial and lateral perforant path-granule cell synapses in the dentate gyrus. Low-frequency stimulation failed to induce long-term depression in the CA1 region of TN-C-deficient animals. Recordings of TBS-induced LTP in the presence of nifedipine, an antagonist of L-type voltage-dependent Ca2+ channels (VDCCs), did not affect LTP in TN-C-deficient mice, but reduced LTP in wild-type mice to the levels seen in mutants. Furthermore, chemical induction of a L-type VDCC-dependent LTP in the CA1 region by application of the K+ channel blocker tetraethylammonium resulted in impaired LTP in TN-C mutants. Thus, reduction in L-type VDCC-mediated signaling appears to mediate the deficits in certain forms of synaptic plasticity in constitutively TN-C-deficient mice.

Perfusion single photon emission computed tomography in a mouse model of neurofibromatosis type 1: towards a biomarker of neurologic deficits.

Neurofibromatosis type 1 (NF1) is a single-gene disorder affecting neurologic function in humans. The NF1+/- mouse model with germline mutation of the NF1 gene presents with deficits in learning, attention, and motor coordination, very similar to NF1 patients. The present study performed brain perfusion single-photon emission computed tomography (SPECT) in NF1+/- mice to identify possible perfusion differences as surrogate marker for altered cerebral activity in NF1. Cerebral perfusion was measured with hexamethyl-propyleneamine oxime (HMPAO) SPECT in NF1+/- mice and their wild-type littermates longitudinally at juvenile age and at young adulthood. Histology and immunohistochemistry were performed to test for structural changes. There was increased HMPAO uptake in NF1 mice in the amygdala at juvenile age, which reduced to normal levels at young adulthood. There was no genotype effect on thalamic HMPAO uptake, which was confirmed by ex vivo measurements of F-18-fluorodeoxyglucose uptake in the thalamus. Morphologic analyses showed no major structural abnormalities. However, there was some evidence of increased density of microglial somata in the amygdala of NF1-deficient mice. In conclusion, there is evidence of increased perfusion and increased density of microglia in juvenile NF1 mice specifically in the amygdala, both of which might be associated with altered synaptic plasticity and, therefore, with cognitive deficits in NF1.

GluK2-mediated excitability within the superficial layers of the entorhinal cortex.

Recent analysis of genetically modified mice deficient in different kainate receptor (KAR) subunits have strongly pointed to a role of the GluK2 subunit, mediating the vulnerability of the brain towards seizures. Research concerning this issue has focused mainly on the hippocampus. However, several studies point to a potential role of other parts of the hippocampal formation, in particular the entorhinal cortex, in the development of epileptic seizures. There is extensive cell death after such seizures in layer III of the medial entorhinal cortex (LIII mEC), making this region of special interest for investigation into related pathological conditions. We therefore characterized KAR mediated currents in LIII mEC pyramidal neurons by several different approaches. Using patch-clamp technique, in combination with glutamate uncaging in horizontal brain slices, we show that LIII mEC neurons exhibit KAR currents. Use of genetically modified mice reveal that these currents are mediated by GluK2 containing KARs. The IV curve indicates the predominant presence of a Ca(2+) impermeable and edited form of the KAR. Finally, we show that GluK2 containing kainate receptors are essential for kainate-induced gamma oscillations within the entorhinal cortex.