Ventral tegmental area astrocytes orchestrate avoidance and approach behavior.

The ventral tegmental area (VTA) is a heterogeneous midbrain structure, containing neurons and astrocytes, that coordinates behaviors by integrating activity from numerous afferents. Within neuron-astrocyte networks, astrocytes control signals from distinct afferents in a circuit-specific manner, but whether this capacity scales up to drive motivated behavior has been undetermined. Using genetic and optical dissection strategies we report that VTA astrocytes tune glutamatergic signaling selectively on local inhibitory neurons to drive a functional circuit for learned avoidance. In this circuit, astrocytes facilitate excitation of VTA GABA neurons to increase inhibition of dopamine neurons, eliciting real-time and learned avoidance behavior that is sufficient to impede expression of preference for reward. Loss of one glutamate transporter (GLT-1) from VTA astrocytes selectively blocks these avoidance behaviors and spares preference for reward. Thus, VTA astrocytes selectively regulate excitation of local GABA neurons to drive a distinct avoidance circuit that opposes approach behavior.

Inhibition of the Agrobacterium tumefaciens TraR quorum-sensing regulator. Interactions with the TraM anti-activator.

The Agrobacterium tumefaciens quorum-sensing transcriptional regulator TraR and its inducing ligand 3-oxo-octanoyl-l-homoserine lactone control conjugal transfer of the tumor-inducing plasmid, the primary virulence factor responsible for crown gall disease of plants. This regulatory system enables A. tumefaciens to express its conjugal transfer regulon preferentially at high population densities. TraR activity is antagonized by a second tumor-inducing plasmid-encoded protein designated TraM. TraM and TraR are thought to form an anti-activation complex that prevents TraR from recognizing its target DNA-binding sites. The formation and inhibitory function of the TraM-TraR anti-activation complex was analyzed using several different assays for protein-protein interaction, including surface plasmon resonance. The TraR-TraM complex forms readily in solution and is extremely stable (K(D) of 1-4 x 10(-9) m). Directed mutational analysis of TraM identified a number of amino acids that play important roles in the inhibition of TraR, clustering in two regions of the protein. Interestingly, several mutants were identified that proficiently bound TraR but were unable to inhibit its activity. This observation suggests a mechanistic separation between the initial assembly of the complex and conversion of TraR to an inactive form.

The hairless gene mutated in congenital hair loss disorders encodes a novel nuclear receptor corepressor.

The mammalian hairless (hr) gene plays a critical role in the maintenance of hair growth. Although the hr gene has been identified, the biochemical function of its encoded protein (Hr) has remained obscure. Here, we show that Hr functions as a transcriptional corepressor for thyroid hormone receptors (TRs). We find that two independent regions of Hr mediate TR binding and that interaction requires a cluster of hydrophobic residues similar to the binding motifs proposed for nuclear receptor corepressors (N-CoR and SMRT). Similarly, we show that Hr binds to the same region of TR as known corepressors. We show that Hr interacts with histone deacetylases (HDACs) and is localized to matrix-associated deacetylase (MAD) bodies, indicating that the mechanism of Hr-mediated repression is likely through associated HDAC activity. Thus, Hr is a component of the corepressor machinery, and despite its lack of sequence identity with previously described corepressors, its mode of action is remarkably conserved. On the basis of its thyroid hormone-inducible and tissue- and developmental-specific expression, Hr likely defines a new class of nuclear receptor corepressors that serve a more specialized role than ubiquitous corepressors. The discovery that Hr is a corepressor provides a molecular basis for specific hair loss syndromes in both humans and mice.

Neuronal expression of synaptotagmin-related gene 1 is regulated by thyroid hormone during cerebellar development.

Thyroid hormone (TH) is essential for proper brain development, acting through nuclear receptors that modulate the expression of specific genes in response to hormone binding. In a screen for genes regulated by TH in the rat cerebellum, we recently identified a novel gene, synaptotagmin-related gene 1 (Srg1). The Srg1 protein is structurally similar to synaptotagmins, a family of proteins involved in regulating neurotransmission. To elucidate a potential role of Srg1 in brain development, we have investigated the developmental and TH-regulated expression of Srg1 in the neonatal rat brain. We show that expression of both Srg1 RNA and protein is detected only in the brain and specifically in neurons. Srg1 mRNA and protein levels increase postnatally, nearing adult levels after the third postnatal week. Neonatal TH deficiency results in a significant reduction and delay in expression of both Srg1 RNA and protein. Using immunohistochemistry, we were able to detect Srg1 protein in numerous brain regions. In the cerebellum, Srg1 protein is localized to the molecular layer, indicating that it is highly expressed in granule cell axons. To further examine Srg1 expression in cerebellar granule cells (CGCs), we used an in vitro cell culture model. In primary cultures of CGCs, Srg1 expression is significantly reduced in the absence of TH. Srg1 mRNA is rapidly upregulated in cultured CGCs, suggesting a direct response to TH. Neuronal and TH-regulated expression of Srg1, together with its localization to neurites, implicates Srg1 as an important component of the program of gene expression induced by TH in the developing brain.

Measurement of bilirubin partition coefficients in bile salt micelle/aqueous buffer solutions by micellar electrokinetic chromatography.

The partition coefficients for the distribution of bilirubin between aqueous phosphateborate buffer and cholic, taurocholic, taurodeoxycholic, and taurochenodeoxycholic micelles have been measured by micellar electrokinetic chromatography at pH 8.5. Determination of the partition coefficients required that the critical micelle concentration and partial specific volumes be determined for each bile salt. Critical micelle concentrations were slightly higher for the trihydroxy bile salts. Partial specific volumes of the bile salt micelles differed very little from each other, and for each bile salt they were constant over the concentration range studied, which was typically from slightly above the critical micelle concentration to 35 mM. Capacity factors were corrected for the effects of applied voltage by extrapolation of the capacity factor to zero applied volts. The free solution mobility of bilirubin, determined in the absence of bile salt, was also corrected for the effects of applied voltage. Plots of extrapolated capacity factor versus phase ratio yield the partition coefficient as the slope of a linear fit to the data. Partition coefficients for bilirubin were significantly higher for dihydroxy bile salts than for trihydroxy bile salts.