Ultrastructural abnormalities in CA1 hippocampus caused by deletion of the actin regulator WAVE-1.

By conveying signals from the small GTPase family of proteins to the Arp2/3 complex, proteins of the WAVE family facilitate actin remodeling. The WAVE-1 isoform is expressed at high levels in brain, where it plays a role in normal synaptic processing, and is implicated in hippocampus-dependent memory retention. We used electron microscopy to determine whether synaptic structure is modified in the hippocampus of WAVE-1 knockout mice, focusing on the neuropil of CA1 stratum radiatum. Mice lacking WAVE-1 exhibited alterations in the morphology of both axon terminals and dendritic spines; the relationship between the synaptic partners was also modified. The abnormal synaptic morphology we observed suggests that signaling through WAVE-1 plays a critical role in establishing normal synaptic architecture in the rodent hippocampus.

Ecto-nucleoside triphosphate diphosphohydrolase 3 in the ventral and lateral hypothalamic area of female rats: morphological characterization and functional implications.

BACKGROUND: Based on its distribution in the brain, ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3) may play a role in the hypothalamic regulation of homeostatic systems, including feeding, sleep-wake behavior and reproduction. To further characterize the morphological attributes of NTPDase3-immunoreactive (IR) hypothalamic structures in the rat brain, here we investigated: 1.) The cellular and subcellular localization of NTPDase3; 2.) The effects of 17beta-estradiol on the expression level of hypothalamic NTPDase3; and 3.) The effects of NTPDase inhibition in hypothalamic synaptosomal preparations. METHODS: Combined light- and electron microscopic analyses were carried out to characterize the cellular and subcellular localization of NTPDase3-immunoreactivity. The effects of estrogen on hypothalamic NTPDase3 expression was studied by western blot technique. Finally, the effects of NTPDase inhibition on mitochondrial respiration were investigated using a Clark-type oxygen electrode. RESULTS: Combined light- and electron microscopic analysis of immunostained hypothalamic slices revealed that NTPDase3-IR is linked to ribosomes and mitochondria, is predominantly present in excitatory axon terminals and in distinct segments of the perikaryal plasma membrane. Immunohistochemical labeling of NTPDase3 and glutamic acid decarboxylase (GAD) indicated that gamma-amino-butyric-acid- (GABA) ergic hypothalamic neurons do not express NTPDase3, further suggesting that in the hypothalamus, NTPDase3 is predominantly present in excitatory neurons. We also investigated whether estrogen influences the expression level of NTPDase3 in the ventrobasal and lateral hypothalamus. A single subcutaneous injection of estrogen differentially increased NTPDase3 expression in the medial and lateral parts of the hypothalamus, indicating that this enzyme likely plays region-specific roles in estrogen-dependent hypothalamic regulatory mechanisms. Determination of mitochondrial respiration rates with and without the inhibition of NTPDases confirmed the presence of NTPDases, including NTPDase3 in neuronal mitochondria and showed that blockade of mitochondrial NTPDase functions decreases state 3 mitochondrial respiration rate and total mitochondrial respiratory capacity. CONCLUSION: Altogether, these results suggest the possibility that NTPDases, among them NTPDase3, may play an estrogen-dependent modulatory role in the regulation of intracellular availability of ATP needed for excitatory neuronal functions including neurotransmission.

Synaptic alpha-dystrobrevin: localization of a short alpha-dystrobrevin isoform in melanin-concentrating hormone neurons of the hypothalamus.

The expression of the two members of the dystrobrevin (DB) family in the adult brain was thought to be highly specific for the two main cell types: alpha-dystrobrevin (alpha-DB) and beta-dystrobrevin (beta-DB) has been identified as glial and neuronal proteins, respectively. In the present work we show that a subset of neurons in the hypothalamus contains alpha-DB. Comparative immunohistochemical studies with two alpha-DB antibodies of different specificity indicate that the neurons contain short alpha-DB isoform(s) alpha-DB-2 and/or alpha-DB-4. Immunoreactive multipolar or spindle-shaped neurons form clusters with bilateral symmetry, localized predominantly in the lateral hypothalamic area, with extensions into the zona incerta and the dorso-medial and ventro-medial hypothalamic region. alpha-DB immunoreactivity was localized in cell processes and at postsynaptic densities, furthermore in the endoplasmic reticulum within the perikarya. alpha-DB-positive neurons are beta-dystrobrevin immunoreactive, but alpha- and beta-DB do not co-localize with their usual molecular anchors like dystrophins or high molecular weight forms of utrophin. Colocalization with nNOS was also not observed. The pattern of alpha-DB immunoreactive neurons gave a perfect colocalization with melanin-concentrating hormone (MCH) neurons throughout the whole region studied. We propose that alpha-DB plays a role in a structure or regulation mechanism unique to MCH-expressing neurons.

Expression of alpha-dystrobrevin in blood-tissue barriers: sub-cellular localisation and molecular characterisation in normal and dystrophic mice.

The alpha- and beta-dystrobrevins (DBs) belong to a family of dystrophin-related and dystrophin-associated proteins that are members of the dystrophin-associated protein complex (DAPC). This complex provides a link between the cytoskeleton and the extracellular matrix or other cells. However, specific functions of the two dystrobrevins remain largely unknown, with alpha-DB being believed to have a role mainly in skeletal muscle. Here, we describe previously unknown expression patterns and the localisation and molecular characteristics of alpha-DB isoforms in non-muscle mouse tissues. We demonstrate a highly specific sub-cellular distribution of alpha-DB in organs forming blood-tissue barriers. We show alpha-DB expression and localisation in testicular Sertoli cells, stomach and respiratory epithelia and provide electron-microscopic evidence for its immunolocalisation in these cells and in the central nervous system. Moreover, we present the molecular characterisation of alpha-DB transcript in these tissues and provide evidence for a distinct heterogeneity of associations between alpha-DB and dystrophins and utrophin in normal and dystrophic non-muscle tissues. Together, our results indicate that alpha-DB, in addition to its role in skeletal muscle, may also be required for the proper function of specific non-muscle tissues and that disruption of DAPC might lead to tissue-blood barrier abnormalities.

Dystrophin splice variants are distinctly localized in the hippocampus.

It has previously been demonstrated that Dp71, the most abundant dystrophin protein in the brain, is mainly localized in the postsynaptic densities. Here we show the localization of Dp71f, one of the splice variants of this protein, within the CA3 region of the hippocampus. Immunopositivity occurs in the postsynaptic density of small asymmetrical axospinous and axodendritic synapses, while it is absent in the postsynaptic densities of the axospinous synapses of the large mossy fiber terminals. Dp71f immunoreactivity was found to be attached to the membranes of the mossy fibers in the stratum lucidum of the CA3 area. In a certain population of thin myelinated axons the protein seems to be present within the axon proper. These data support the notion of a physiological role of Dp71f distinct from other dystrophin isoforms present in the central nervous system.