Mutations of the gene coding for PAK3 (p21-activated kinase 3) are associated with X-linked, nonsyndromic forms of mental retardation (MRX) in which the only distinctive clinical feature is the cognitive deficit. The mechanisms through which PAK3 mutation produces the mental handicap remain unclear, although an involvement in the mechanisms that regulate the formation or plasticity of synaptic networks has been proposed. Here we show, using a transient transfection approach, that antisense and small interfering RNA-mediated suppression of PAK3 or expression of a dominant-negative PAK3 carrying the human MRX30 mutation in rat hippocampal organotypic slice cultures results in the formation of abnormally elongated dendritic spines and filopodia-like protrusions and a decrease in mature spine synapses. Ultrastructural analysis of the changes induced by expression of PAK3 carrying the MRX30 mutation reveals that many elongated spines fail to express postsynaptic densities or contact presynaptic terminals. These defects are associated with a reduced spontaneous activity, altered expression of AMPA-type glutamate receptors, and defective long-term potentiation. Together, these data identify PAK3 as a key regulator of synapse formation and plasticity in the hippocampus and support interpretations that these defects might contribute to the cognitive deficits underlying this form of mental retardation.

We investigated rearrangements of connectivity between hippocampal mossy fibers and CA3 pyramidal neurons. We found that mossy fibers establish 10-15 local terminal arborization complexes (LMT-Cs) in CA3, which exhibit major differences in size and divergence in adult mice. LMT-Cs exhibited two types of long-term rearrangements in connectivity in the adult: progressive expansion of LMT-C subsets along individual dendrites throughout life, and pronounced increases in LMT-C complexities in response to an enriched environment. In organotypic slice cultures, subsets of LMT-Cs also rearranged extensively and grew over weeks and months, altering the strength of preexisting connectivity, and establishing or dismantling connections with pyramidal neurons. Differences in LMT-C plasticity reflected properties of individual LMT-Cs, not mossy fibers. LMT-C maintenance and growth were regulated by spiking activity, mGluR2-sensitive transmitter release from LMTs, and PKC. Thus, subsets of terminal arborization complexes by mossy fibers rearrange their local connectivities in response to experience and age throughout life.

Cerebral ischemia is a major cause of brain dysfunction. Using a model of delayed death induced by a brief, transient oxygen and glucose deprivation, we studied here how this affected the structural organization of hippocampal synaptic networks. We report that brief anoxic-hypoglycemic episodes rapidly modified the structure of synapses. This was characterized, at the electron microscopic level, by a transient increase in the proportion of perforated synapses, followed after 2 hr by an increase in images of multiple synapse boutons. These changes were considerable because 10-20% of all synapses were affected. This structural remodeling was correlated by three kinds of modifications observed using two-photon confocal microscopy: the growth of filopodia, occurring shortly (5-20 min) after anoxia-hypoglycemia, enlargements of existing spines, and formation of new spines, both seen mainly 20-60 min after the insult. All of these structural changes were calcium and NMDA receptor dependent and thus reproduced, to a larger scale, those associated with synaptic plasticity. Concomitantly and related to the severity of anoxia-hypoglycemia, we could also observe spine loss and images of spine, dendrite, or presynaptic terminal swellings that evolved up to membrane disruption. These changes were also calcium dependent and reduced by NMDA receptor antagonists. Thus, short anoxic-hypoglycemic episodes, through NMDA receptor activation and calcium influx, resulted in a profound structural remodeling of synaptic networks, through growth, formation, and elimination of spines and synapses.

Trafficking of AMPA-type glutamate receptors (AMPAR) between endosomes and the postsynaptic plasma membrane of neurons plays a central role in the control of synaptic strength associated with learning and memory. The molecular mechanisms of its regulation remain poorly understood, however. Here we show by biochemical and atomic force microscopy analyses that NEEP21, a neuronal endosomal protein necessary for receptor recycling including AMPAR, is associated with the scaffolding protein GRIP1 and the AMPAR subunit GluR2. Moreover, the interaction between NEEP21 and GRIP1 is regulated by neuronal activity. Expression of a NEEP21 fragment containing the GRIP1-binding site decreases surface GluR2 levels and delays recycling of internalized GluR2, which accumulates in early endosomes and lysosomes. Infusion of this fragment into pyramidal neurons of hippocampal slices induces inward rectification of AMPAR-mediated synaptic responses, suggesting decreased GluR2 expression at synapses. These results indicate that NEEP21-GRIP1 binding is crucial for GluR2-AMPAR sorting through endosomes and their recruitment to the plasma membrane, providing a first molecular mechanism to differentially regulate AMPAR subunit cycling in internal compartments.

Our current understanding of the mechanisms of information processing and storage in the brain, based on the concept proposed more than fifty years ago by D. Hebb, is that a key role is played by changes in synaptic efficacy induced by coincident pre- and postsynaptic activity. Decades of studies of the properties of long-term potentiation (LTP) have shown that this form of plasticity adequately fulfills these requirements and is likely to contribute to several models of learning and memory. Recent analyses of the molecular events implicated in LTP are consistent with the view that modifications of receptor properties or insertion of new receptors account for the potentiation of synaptic transmission. These experiments, however, have also uncovered an unexpected structural plasticity of synapses. Dendritic spines appear to be dynamic structures that can be formed, modified in their shape or eliminated under the influence of activity. Furthermore, recent studies suggest that LTP, in addition to changes in synaptic function, is also associated with mechanisms of synaptogenesis. We review here the evidence pointing to this activity-dependent remodeling and discuss the possible role of this structural plasticity for synaptic potentiation, learning and memory.