Jozsef Zoltan Kiss

PSA–NCAM Is Required for Activity-Induced Synaptic Plasticity

Hippocampal organotypic slice cultures maintained 10-20 days in vitro express a high level of the polysialylated embryonic form of neural cell adhesion molecule (NCAM) (PSA-NCAM). Treatment of the cultures with endoneuraminidase-N selectively removed polysialic acid (PSA) from NCAM and completely prevented induction of long-term potentiation (LTP) and long-term depression (LTD) without affecting cellular or synaptic parameters. Similarly, slices prepared from transgenic mice lacking the NCAM gene exhibited a decaying LTP. No inhibition of N-methyl-D-aspartic acid receptor-dependent synaptic responses was detected. Washout of the enzyme resulted in reexpression of PSA immunoreactivity which correlated with a complete recovery of LTP and LTD. This reexpression was blocked by TTX and low calcium and enhanced by bicuculline. Taken together, these results indicate that neuronal activity regulates the expression of PSA-NCAM at the synapse and that this expression is required for the induction of synaptic plasticity.

Extracellular proteolysis in the adult murine brain.

Plasminogen activators are important mediators of extracellular metabolism. In the nervous system, plasminogen activators are thought to be involved in the remodeling events required for cell migration during development and regeneration. We have now explored the expression of the plasminogen activator/plasmin system in the adult murine central nervous system. Tissue-type plasminogen activator is synthesized by neurons of most brain regions, while prominent tissue-type plasminogen activator-catalyzed proteolysis is restricted to discrete areas, in particular within the hippocampus and hypothalamus. Our observations indicate that tissue-type plasminogen activator-catalyzed proteolysis in neural tissues is not limited to ontogeny, but may also contribute to adult central nervous system physiology, for instance by influencing neuronal plasticity and synaptic reorganization. The identification of an extracellular proteolytic system active in the adult central nervous system may also help gain insights into the pathogeny of neurodegenerative disorders associated with extracellular protein deposition.

VEGF is a chemoattractant for FGF-2-stimulated neural progenitors

Mmigration of undifferentiated neural progenitors is critical for the development and repair of the nervous system. However, the mechanisms and factors that regulate migration are not well understood. Here, we show that vascular endothelial growth factor (VEGF)-A, a major angiogenic factor, guides the directed migration of neural progenitors that do not display antigenic markers for neuron- or glia-restricted precursor cells. We demonstrate that progenitor cells express both VEGF receptor (VEGFR) 1 and VEGFR2, but signaling through VEGFR2 specifically mediates the chemotactic effect of VEGF. The expression of VEGFRs and the chemotaxis of progenitors in response to VEGF require the presence of fibroblast growth factor 2. These results demonstrate that VEGF is an attractive guidance cue for the migration of undifferentiated neural progenitors and offer a mechanistic link between neurogenesis and angiogenesis in the nervous system.

Stress-induced changes in messenger RNA levels of N-methyl-d-aspartate and AMPA receptor subunits in selected regions of the rat hippocampus and hypothalamus

The postsynaptic AMPA/kainate and N-methyl-D-aspartate-selective glutamate receptors are formed by several different subunits and the overall subunit composition of the receptor appears to determine its physiological and pharmacological properties. Although glutamatergic mechanisms have been implicated in various forms of hippocampal stress responses, the impact of stress on glutamate receptor subunit composition has not yet been elucidated. We have used cell-by-cell quantitative in situ hybridization to assess stress-induced changes in transcript levels of N-methyl-D-aspartate and AMPA receptor subunit genes in subdivisions of the rat hippocampus and hypothalamus that are implicated in the stress response. We found that 24 h after a single immobilization stress there was a significant increase in the cellular level of NR1 subunit messenger RNA (about 35-45% above control values) in hippocampal CA3 and CA1 pyramidal cells as well as in neurons of the hypothalamic supraoptic and paraventricular nuclei. Moreover, in the CA3 area we have detected a concomitant increase (50% above controls) in the level of NR2B subunit messenger RNA, while the expression of NR2A subunit gene did not change after stress. Stress induced a selective decrease in the level of AMPA receptor subunit glutamate receptor A messenger RNA in neurons of both the CA3 and CA1 areas (18 and 24%, respectively, below control values). These results suggest that the regulation of specific subunit messenger RNAs of the N-methyl-D-aspartate and AMPA receptors may be involved in altered hippocampal and hypothalamic responsiveness to glutamate and thus could play a critical role in stress-induced changes in their function.

PSA‐NCAM modulates BDNF‐dependent survival and differentiation of cortical neurons

We show that the loss or inactivation of the polysialic acid (PSA) tail of neural cell adhesion molecule (NCAM) on rat cortical neurons in culture leads to reduced differentiation and survival. The mechanism by which this negative effect is mediated appears to involve the neuronal response to brain‐derived neurotrophic factor (BDNF): (i) in the absence of PSA or in the presence of excess free PSA added to the culture medium, BDNF‐induced cell signalling is reduced; (ii) the addition of exogenous BDNF to the medium reverses the effect of PSA loss or inactivation. These data suggest that PSA‐NCAM, previously shown to modulate cell migration and plasticity, is needed for an adequate sensitivity of neurons to BDNF.

Contribution of the neural cell adhesion molecule to neuronal and synaptic plasticity.

The neural cell adhesion molecule (NCAM) and its polysialylated form PSA-NCAM contribute to many aspects of the development and plasticity of the central nervous system. This includes mechanisms of cell differentiation and migration, neurite outgrowth, establishment of specific patterns of synaptic connections, synaptic plasticity and long-term potentiation. How NCAM and PSA-NCAM contribute to regulate all these different mechanisms remains essentially unknown. Adhesive properties appear to be important, but recent studies also point to possible interactions between NCAM and PSA-NCAM with intracellular signalling cascades that are essential to biological functions. Some of these mechanisms are discussed and a hypothesis is proposed based on the existence of cross-talk between these molecules and signalling pathways mediated by growth factors.

Activity-dependent mobilization of the adhesion molecule polysialic NCAM to the cell surface of neurons and endocrine cells.

The alpha‐2,8‐linked sialic acid polymer (PSA) on the neural cell adhesion molecule (NCAM) is an important regulator of cell surface interactions. We have examined the translocation of PSA‐NCAM to the surface of cultured cortical neurons and insulin secreting beta cells under different conditions of cell activity. Endoneuraminidase N, an enzyme that specifically cleaves PSA chains, was used to remove pre‐existing PSA from the plasma membrane and the re‐expression of the molecule was monitored by immunocytochemistry. Punctate PSA immunostaining was restored on the surface of 68% of neurons within 1 h. This recovery was almost completely prevented by tetrodotoxin, suggesting that spontaneous electrical activity is required. K+ depolarization (50 mM) allowed recovery of PSA surface staining in the presence of tetrodotoxin and this effect required the presence of extracellular Ca2+. Rapid redistribution of PSA‐NCAM to the surface of beta cells was observed under conditions that stimulate insulin secretion. Ca2+ channel inhibition decreased both PSA‐NCAM expression and insulin secretion to control, non‐stimulated levels. Finally, subcellular fractionation of an insulin‐secreting cell line showed that the secretory vesicle fraction is highly enriched in PSA‐NCAM. These results suggest that PSA‐NCAM can be translocated to the cell surface via regulated exocytosis. Taken together, our results provide unprecedented evidence linking cell activity and PSA‐NCAM expression, and suggest a mechanism for rapid modulation of cell surface interactions.

Serotoninergic endings on VIP-neurons in the suprachiasmatic nucleus and on ACTH-neurons in the arcuate nucleus of the rat hypothalamus. A combination of high resolution autoradiography and electron microscopic immunocytochemistry

A combination of immunocytochemistry with transmitter specific autoradiography at the electron microscopic level was used to identify vasoactive intestinal polypeptide (VIP)- or ACTH-containing neurons and serotoninergic nerve terminals in the suprachiasmatic (SCN) and arcuate nuclei (AN). We observed nerve terminals showing selective uptake of [3H]serotonin forming synaptic contact with perikarya or dendrites where the postsynaptic structures exhibited VIP-like immunoreactivity in the SCN and ACTH-like immunoreactivity in the AN. The findings provide ultrastructural evidence that serotoninergic nerve fibers terminate on VIP-neurons in the SCN and on ACTH-neurons in the AN.

The role of neural cell adhesion molecules in plasticity and repair

Repair and functional recovery after brain injury critically depends on structural and functional plasticity of preserved neuronal networks. A striking feature of brain structures where tissue reorganization and plasticity occur is a strong expression of the polysialylated neural cell adhesion molecule (PSA-NCAM). An important role of this molecule in various aspects of neuronal and synaptic plasticity has been revealed by many studies. Recently, a new mechanism has been elucidated whereby PSA-NCAM may contribute to signalling mediated by the neurotrophic factor BDNF, thereby sensitizing neurons to this growth factor. This mechanism was shown to be important for activity-induced synaptic plasticity and for the survival and differentiation of cortical neurons. A cross-talk between these molecules may, thus, reveal a key factor for properties of structural plasticity and in particular could mediate the activity-dependent aspects of synaptic network remodeling. Animal models have been developed to assess the role of these molecules in functional recovery after lesions.

Demonstration of serotoninergic axons terminating on luteinizing hormone-releasing hormone neurons in the preoptic area of the rat using a combination of immunocytochemistry and high resolution autoradiography

The synaptic relationship between serotoninergic terminals and luteinizing hormone-releasing hormone-containing neurons was investigated in the medial preoptic area using a combined technique. Axon terminals selectively taking up 5-[3H]hydroxytryptamine were labelled autoradiographically and luteinizing hormone-releasing hormone-containing neuronal elements were identified by means of immunocytochemistry. Synaptic contacts were observed between tritiated 5-hydroxytryptamine-labelled boutons and luteinizing hormone-releasing hormone-immunoreactive dendrites. About 5% of the boutons which formed synapses with luteinizing hormone-releasing hormone-immunoreactive dendrites were found to be labelled by the tritiated indolamine. Luteinizing hormone-releasing hormone-immunoreactive axon terminals occurred as presynaptic elements in contact with unidentified dendritic spines, shafts or perikarya. These observations provide morphological basis for the idea that 5-hydroxytryptamine-containing neurons can act directly on luteinizing hormone-releasing hormone release. Further, they support the assumption that luteinizing hormone-releasing hormone is not only a neurohormone but may also function as a neurotransmitter or neuromodulator.