Newborn neurons in the dentate gyrus of the adult hippocampus rely upon cAMP response element binding protein (CREB) signaling for their differentiation into mature granule cells and their integration into the dentate network. Among its many targets, the transcription factor CREB activates expression of a gene locus that produces two microRNAs, miR-132 and miR-212. In cultured cortical and hippocampal neurons, miR-132 functions downstream from CREB to mediate activity-dependent dendritic growth and spine formation in response to a variety of signaling pathways. To investigate whether miR-132 and/or miR-212 contribute to the maturation of dendrites in newborn neurons in the adult hippocampus, we inserted LoxP sites surrounding the miR-212/132 locus and specifically targeted its deletion by stereotactically injecting a retrovirus expressing Cre recombinase. Deletion of the miR-212/132 locus caused a dramatic decrease in dendrite length, arborization, and spine density. The miR-212/132 locus may express up to four distinct microRNAs, miR-132 and -212 and their reverse strands miR-132* and -212*. Using ratiometric microRNA sensors, we determined that miR-132 is the predominantly active product in hippocampal neurons. We conclude that miR-132 is required for normal dendrite maturation in newborn neurons in the adult hippocampus and suggest that this microRNA also may participate in other examples of CREB-mediated signaling.microRNA-212 | neurogenesis | plasticity | learning

Excitatory amino acid transporters (EAATs) function as both substrate transporters and ligand-gated anion channels. Characterization of the transporter's general topology is the first requisite step in defining the structural bases for these distinct activities. While the first six hydrophobic domains can be readily modeled as conventional transmembrane segments, the organization of the C-terminal hydrophobic domains, which have been implicated in both substrate and ion interactions, has been controversial. Here, we report the results of a comprehensive evaluation of the C-terminal topology of EAAT1 determined by the chemical modification of introduced cysteine residues. Our data support a model in which two membrane-spanning domains flank a central region that is highly accessible to the extracellular milieu and contains at least one reentrant loop domain.

The rapid re-uptake of extracellular glutamate mediated by a family of high-affinity glutamate transporter proteins is essential to continued glutamatergic signaling and neuronal viability, but the contributions of individual transporter subtypes toward cellular physiology are poorly understood. Because the physiology of glutamate transport in the salamander retina has been well described, we have examined the expression and function of glutamate transporter subtypes in this preparation. cDNAs encoding five distinct salamander excitatory amino acid transporter (sEAAT) subtypes were isolated, and their molecular properties and distributions of expression were compared. We report evidence that at least four distinct sEAAT subtypes are expressed in glial (Mü ller) cells. In addition, four of the five transporter subtypes are localized in neurons throughout the retina. The brightest immunostaining was seen in the synaptic regions of the inner and outer plexiform layers and in the outer nuclear layer. Using electrophysiological measurements in the Xenopus oocyte expression system, we also examined the pharmacology and ionic dependence of the four expressing transporter subtypes that make it possible to distinguish, on the basis of functional behavior, among the various subtypes. Although no simple correlation between transporter subtype and retinal cell physiology can be made, the diverse population of sEAAT transporter subtypes with unique localization and functional properties indicates that glutamate transporters play a wide variety of roles in retinal function and are likely to underlie both the uptake of glutamate by Mü ller cells and the glutamateelicited chloride conductance involved in signal transduction by photoreceptors and bipolar cells. Key words: glutamate transporter; uptake; amino acid; retina; chloride; molecular cloning; photoreceptors; Mü ller cells; bipolar cells; immunohistochemistryGlutamate is the predominant excitatory neurotransmitter in the vertebrate retina and is released from photoreceptors, bipolar cells, and ganglion cells (for review, see Massey, 1990). Termination of glutamatergic synaptic transmission requires uptake of glutamate by means of plasma membrane glutamate transporters, which may be present presynaptically and postsynaptically as well as in neighboring glial cells. These transporters are of particular importance in the retina, where glutamate is released continuously and at a high rate in the dark.Electrophysiological studies performed primarily in the salamander retina have demonstrated the existence of glutamate transporters in retinal glial (Müller) cells (Brew and Attwell,