Acute psychotropic, autonomic, and endocrine effects of 5,6-methylenedioxy-2-aminoindane (MDAI) compared with 3,4-methylenedioxymethamphetamine (MDMA) in human volunteers: A self-administration study.

The acute psychoactive, autonomic, and endocrine effects of the new psychoactive substance (NPS) 5,6-methylenedioxy-2-aminoindane (MDAI; 3.0 mg/kg, range 180-228 mg) were investigated in six healthy volunteers (four males, two females) in a non-blinded fashion without placebo. Subjective, cardiovascular, and endocrine responses were compared with two different doses of 3,4-methylenedioxymethamphetamine (MDMA) (75 mg and 125 mg) described in previously published placebo-controlled studies, which used identical outcome measures including Visual Analogue Scales (VAS), the Adjective Mood Rating Scale (AMRS), and the 5 Dimensions of Altered States of Consciousness (5D-ASC) scale. MDAI was well tolerated and produced subjective effects comparable with those of 125 mg MDMA. MDAI increased blood pressure similar to 125 mg MDMA but did not increase heart rate or body temperature. MDAI increased cortisol and prolactin levels and could be detected in serum about 20 min post ingestion and remained detectable at least for 4 days. In urine, MDAI was detectable over a period of at least 6 days. Further clinical investigations are warranted to assess whether MDAI could serve as drug with medicinal properties.

TGF-ß2 Regulates Transcription of the K+/Cl- Cotransporter 2 (KCC2) in Immature Neurons and Its Phosphorylation at T1007 in Differentiated Neurons.

KCC2 mediates extrusion of K+ and Cl- and assuresthe developmental "switch" in GABA function during neuronal maturation. However, the molecular mechanisms underlying KCC2 regulation are not fully elucidated. We investigated the impact of transforming growth factor beta 2 (TGF-ß2) on KCC2 during neuronal maturation using quantitative RT-PCR, immunoblotting, immunofluorescence and chromatin immunoprecipitation in primary mouse hippocampal neurons and brain tissue from Tgf-ß2-deficient mice. Inhibition of TGF-ß/activin signaling downregulates Kcc2 transcript in immature neurons. In the forebrain of Tgf-ß2-/- mice, expression of Kcc2, transcription factor Ap2ß and KCC2 protein is downregulated. AP2ß binds to Kcc2 promoter, a binding absent in Tgf-ß2-/-. In hindbrain/brainstem tissue of Tgf-ß2-/- mice, KCC2 phosphorylation at T1007 is increased and approximately half of pre-Bötzinger-complex neurons lack membrane KCC2 phenotypes rescued through exogenous TGF-ß2. These results demonstrate that TGF-ß2 regulates KCC2 transcription in immature neurons, possibly acting upstream of AP2ß, and contributes to the developmental dephosphorylation of KCC2 at T1007. The present work suggests multiple and divergent roles for TGF-ß2 on KCC2 during neuronal maturation and provides novel mechanistic insights for TGF-ß2-mediated regulation of KCC2 gene expression, posttranslational modification and surface expression. We propose TGF-ß2 as a major regulator of KCC2 with putative implications for pathophysiological conditions.

The membrane trafficking and functionality of the K+-Cl- co-transporter KCC2 is regulated by TGF-ß2.

Functional activation of the neuronal K(+)-Cl(-) co-transporter KCC2 (also known as SLC12A5) is a prerequisite for shifting GABAA responses from depolarizing to hyperpolarizing during development. Here, we introduce transforming growth factor ß2 (TGF-ß2) as a new regulator of KCC2 membrane trafficking and functional activation. TGF-ß2 controls membrane trafficking, surface expression and activity of KCC2 in developing and mature mouse primary hippocampal neurons, as determined by immunoblotting, immunofluorescence, biotinylation of surface proteins and KCC2-mediated Cl(-) extrusion. We also identify the signaling pathway from TGF-ß2 to cAMP-response-element-binding protein (CREB) and Ras-associated binding protein 11b (Rab11b) as the underlying mechanism for TGF-ß2-mediated KCC2 trafficking and functional activation. TGF-ß2 increases colocalization and interaction of KCC2 with Rab11b, as determined by 3D stimulated emission depletion (STED) microscopy and co-immunoprecipitation, respectively, induces CREB phosphorylation, and enhances Rab11b gene expression. Loss of function of either CREB1 or Rab11b suppressed TGF-ß2-dependent KCC2 trafficking, surface expression and functionality. Thus, TGF-ß2 is a new regulatory factor for KCC2 functional activation and membrane trafficking, and a putative indispensable molecular determinant for the developmental shift of GABAergic transmission.

Variants of the electrogenic sodium bicarbonate cotransporter 1 (NBCe1) in mouse hippocampal neurons are regulated by extracellular pH changes: evidence for a Rab8a-dependent mechanism.

Changes in extracellular pH are common events in both pathological conditions and during normal brain function. In organs other than the brain, cells may respond to pH changes by trafficking of acid-base transporters. However, regulation of neuronal acid-base transporters during pH shifts is not understood. The aim of this study was to investigate regulatory mechanisms of the variants of the electrogenic sodium/bicarbonate cotransporter 1, NBCe1-A and NBCe1-B/C, in neurons following changes of extracellular pH. Therefore, primary mouse hippocampal neurons were exposed to extracellular acidosis or alkalosis. We show that acid-base changes regulated trafficking and membrane expression of neuronal NBCe1 but the underlying molecular cues were distinct for individual NBCe1 variants. Following extracellular acidosis NBCe1-A was recruited from intracellular pools to the plasma membrane, followed by increased membrane expression, whereas NBCe1-B/C was retrieved from the membrane. Extracellular alkalosis had no impact on NBCe1-A, but caused translocation of NBCe1-B/C toward the dendrites. We also show that acidosis-induced NBCe1-A, but not NBCe1-B/C, trafficking is mediated by Rab8a. Rab8a is expressed in hippocampal neurons, co-localizes, and interacts with NBCe1-A. Loss-of-function of Rab8a using specific siRNA prevented acidosis-induced redistribution of NBCe1-A. These data propose opposite recruitment pattern for NBCe1 variants in neurons following extracellular acid-base changes, implicating distinct physiological functions of individual NBCe1 variants, and introduce Rab8a as a novel molecular determinant and crucial mediator of acidosis-induced NBCe1 trafficking in neurons.