An accurate method for the quantification of cytochrome C oxidase in tissue sections.

Cytochrome oxidase (COX) is the enzyme that constitutes the last step of the mitochondrial electron transport chain for the production of ATP. Measurement of COX activity can be achieved by histochemistry, thus providing information about the metabolic status of the brain. Brain regions with high metabolism will present high COX activity in histochemistry assays and vice versa.

A new use for long-term frozen brain tissue: golgi impregnation.

The study of dendritic spine shape and number has become a standard in the analysis of synaptic transmission anomalies since a considerable number of neuropsychiatric and neurological diseases have their foundation in alterations in these structures. One of the best ways to study possible alterations of dendritic spines is the use of Golgi impregnation. Although usually the Golgi method implies the use of fresh or fixed tissue, here we report the use of Golgi-Cox for the staining of human and animal brain tissue kept frozen for long periods of time.

Acute regulation of epithelial sodium channel by anionic phospholipids.

Anionic phospholipids such as phosphatidylinositol 4,5-bisphosphate (PIP(2)) and phosphatidylinositol 3,4,5-trisphosphate (PIP(3)) are normally located in the inner leaflet of the plasma membrane, where these anionic phospholipids can regulate transmembrane proteins, including ion channels and transporters. Recent work has demonstrated that (1) ATP inhibits the renal epithelial sodium channel (ENaC) via a phospholipase C-dependent pathway that reduces PIP(2), (2) aldosterone stimulates ENaC via phosphoinositide 3-kinase, and (3) PIP(2) and PIP(3) regulate ENaC. Several lines of evidence show that ATP stimulation of purinergic P2Y receptors hydrolyzes PIP(2) and that aldosterone stimulation of steroid receptors induces PIP(3) formation.

cAMP-induced stellation in primary astrocyte cultures with regional heterogeneity.

It is well known that increased cAMP levels in cultured astrocytes can convert flat polygonal shaped astrocytes into process-bearing, stellate astrocytes. In this study, we have examined the possible existence of astrocyte regional heterogeneity in morphological changes in response to cAMP stimulation. Primary astrocyte cultures were prepared from six different regions of neonatal rat brains, including cerebral cortex, hippocampus, brain stem, mid brain, cerebellum, and hypothalamus.

Regulation of Na+ channel beta 1 and beta 2 subunit mRNA levels in cultured rat astrocytes.

Using quantitative competitive reverse transcription-polymerase chain reaction (RT-PCR), we found an increased level of Na+ channel beta 1 (Na beta 1) mRNAs in spinal cord astrocytes and in the B50 neuroblastoma cell line after exposure to 1 mM dibutyryl cAMP. In contrast, the calcium ionophore (1 microM A23187) did not affect Na beta 1 mRNA levels in these cells. Further, we amplified full length coding region of Na+ channel beta 2 (Na beta 2) mRNA from rat optic nerve and cultured astrocytes using RT-PCR.