[Zbtb33 Gene Knockout Changes Transcription of the Fgf9, Fgfr3, с-Мус and FoxG1 Genes in the Developing Mouse Brain].

The transcription factor KAISO is important for proper development of animal embryos. In the cell, KAISO regulates cell division and apoptosis. KAISO is abundant in the central nervous system. Here we describe the effects of Zbtb33 gene knockout on the transcription of several genes that regulate the development of the central nervous system, including Fgf9, Fgfr3, Sox9, Sox2, c-Myc, NeuroD1 and FoxG1. These genes are related to the Wnt/ß-catenin signaling pathway, which is closely connected to KAISO. Hippocampal, frontal cortical, and striatal tissue from C57BL/6j mice with a knockout in the Zbtb33 gene encoding KAISO (ZBTB33-) and wild-type mice (ZBTB33+) were collected and profiled at different stages of development. Age-dependent and region-specific differences in the mRNA levels of the Fgf9, Fgfr3, c-Myc, FoxG1 genes in the developing brain of ZBTB33- and ZBTB33+ mice were described and discussed.

Inhibitor of Striatal-Enriched Protein Tyrosine Phosphatase, 8-(Trifluoromethyl)-1,2,3,4,5-Benzopentathiepin-6-Amine hydrochloride (TC-2153), Produces Antidepressant-Like Effect and Decreases Functional Activity and Protein Level of 5-HT2A Receptor in the Brain.

The serotoninergic 5-HT2A receptor is involved in the mechanism of depression and antidepressant drugs action. Earlier we showed that striatal-enriched protein tyrosine phosphatase (STEP) inhibitor - 8-(trifluoromethyl)-1,2,3,4,5-benzopentathiepin-6-amine hydrochloride (TC-2153) affects both the brain serotoninergic system and the brain-derived neurotropic factor that are known to be involved in the psychopathology of depression. In the present study we investigated the effects of chronic TC-2153 administration on behavior in the standard battery of tests as well as the effects of acute and chronic TC-2153 treatment on the brain 5-HT2A receptors in mice. We obtained a prominent antidepressant-like effect of chronic TC-2153 treatment in the forced swim test without any adverse side effects on locomotor activity, anxiety, exploration, motor skill and obsessive-compulsive-like behavior. Moreover, both acute and chronic TC-2153 administration inhibited the functional activity of 5-HT2A receptors estimated by the number of 2,5-dimethoxy-4-iodoamphetamine (DOI, agonist of 5-HT2A receptors)-induced head-twitches. TC-2153 treatment also attenuated the DOI-induced c-fos expression in cortical and hippocampal neurons and reduced the 5-HT2A receptor protein level in the hippocampus and frontal cortex, but not in the striatum. Taken together, our combined data demonstrate that the antidepressant effect of STEP inhibitor TC-2153 could be mediated by its inhibitory properties towards the 5-HT2A receptor-mediated signaling.

Effects of a compound from the group of substituted thiadiazines with hypothermia inducing properties on brain metabolism in rats, a study in vivo and in vitro.

The aim of the present study was to examine how administration of a compound of 1,3,4- thiadiazine class 2-morpholino-5-phenyl-6H-1,3,4-thiadiazine, hydrobromide (L-17) with hypothermia inducing properties affects the brain metabolism. The mechanism by which L-17 induces hypothermia is unknown; it may involve hypothalamic central thermoregulation as well as act via inhibition of energy metabolism. We tested the hypothesis that L-17 may induce hypothermia by directly inhibiting energy metabolism. The study in vivo was carried out on Sprague-Dawley adult rats. Two doses of L-17 were administered (190 mg/kg and 760 mg/kg). Brain metabolites were analyzed in control and treated groups using magnetic resonance spectroscopy, along with blood flow rate measurements in carotid arteries and body temperature measurements. Further in vitro studies on primary cultures from rat hippocampus were carried out to perform a mitochondria function test of L-17 pre-incubation (100 µM, 30 min). Analysis of brain metabolites showed no significant changes in 190 mg/kg treated group along with a significant reduction in body temperature by 1.5°C. However, administration of L-17 in higher dose 760 mg/kg provoked changes in brain metabolites indicative of neurotoxicity as well as reduction in carotid arteries flow rate. In addition, a balance change of excitatory and inhibitory neurotransmitters was observed. The L-17 pre-incubation with cell primary cultures from rat brain showed no significant changes in mitochondrial function. The results obtained in the study indicate that acute administration of L-17 190 mg/kg in rats induces mild hypothermia with no adverse effects onto brain metabolism.

Identification of new M23A mRNA of mouse aquaporin-4 expressed in brain, liver, and kidney.

Aquaporins (AQPs) belong to a transmembrane protein family of water channels that are permeable to water by the osmotic gradient. There are two isoforms of mouse AQP4 - M1 and M23. Their balance in the cell determines water permeability of the plasma membrane. These two isoforms are encoded by three mRNAs: M1 isoform is encoded by M1 mRNA and M23 isoform is encoded by M23 and M23X mRNAs. Here we found a new fourth mRNA of mouse AQP4 - M23A mRNA. The start of transcription is different for M23A mRNA from all the known AQP4 mRNAs. The 5'-untranslated region (5'-UTR) of M23A mRNA is encoded by four new exons (A, B, C, and D), which are located in the 5' region from exon-0 of the AQP4 gene. Alternative splicing between the exons-A, -B, -C, and -D leads to formation of multiple variants of M23A mRNA. We cloned six of these variants, all of which code full length M23 isoform of AQP4. Using RT-PCR we detected tissue-specific expression of the new M23A and already known M23, M23X, and M1 mRNAs. The M23A mRNA is expressed mostly in kidney, liver, and brain. Analysis of mRNA 5'-UTR structure showed low translation efficacy for M1 mRNA in comparison with high translation efficacy for M23A, M23X, and M23 mRNAs. We propose that AQP4 expression is controlled tissue-specifically by independent promoters. Thus multiple AQP4 mRNAs may allow long-term regulation of the balance between M1 and M23 AQP4 isoforms in the cell and thus water permeability of the plasma membrane.

Functional and molecular interactions between aquaporins and Na,K-ATPase.

The water channel aquaporin 4 (AQP4) is abundantly expressed in astrocytes and provides a mechanism by which water permeability of the plasma membrane can be regulated. Astrocytes play a key role in the clearance of both potassium (K(+)) and glutamate released during neuronal activity. Emerging evidence suggests that AQP4 facilitates K(+) clearance by astrocytes and contributes to recovery of neuronal excitability. Here we report that AQP4 can assemble with its regulator metabotropic glutamate receptor 5 (mGluR5) and with Na,K-ATPase; the enzyme responsible for active K(+) transport and for establishing the electrochemical gradient across the cell plasma membrane. We have, by use of pull down assays in rat brain tissue, identified the segment in the AQP4 NH(2)-terminus containing the amino acid residues 23-32 as the site for interaction with Na,K-ATPase catalytic subunit and with mGluR5. Mutagenesis studies revealed that the AQP4 amino acids K27 and W30 are of key importance for interaction with both Na,K-ATPase and mGluR5. To confirm that interaction also occurs within intact cells, we have performed fluorescence resonance energy transfer (FRET) studies in primary astrocytes derived from rat striatum. The results indicate close proximity of wild type AQP4 and Na,K-ATPase in the plasma membrane of rat astrocytes. FRET efficiencies observed with the mutants AQP4 K27A and AQP4 W30A were significantly lower, highlighting the importance of these residues for the interaction between AQP4 and Na,K-ATPase. We conclude that AQP4/Na,K-ATPase/mGluR5 can form a macromolecular complex/transporting microdomain in astrocytes. This complex may be of functional importance for the regulation of water and K(+) homeostasis in the brain, as well as for neuron-astrocyte metabolic crosstalk.