Live-cell three-dimensional single-molecule tracking reveals modulation of enhancer dynamics by NuRD.

To understand how the nucleosome remodeling and deacetylase (NuRD) complex regulates enhancers and enhancer-promoter interactions, we have developed an approach to segment and extract key biophysical parameters from live-cell three-dimensional single-molecule trajectories. Unexpectedly, this has revealed that NuRD binds to chromatin for minutes, decompacts chromatin structure and increases enhancer dynamics. We also uncovered a rare fast-diffusing state of enhancers and found that NuRD restricts the time spent in this state. Hi-C and Cut&Run experiments revealed that NuRD modulates enhancer-promoter interactions in active chromatin, allowing them to contact each other over longer distances. Furthermore, NuRD leads to a marked redistribution of CTCF and, in particular, cohesin. We propose that NuRD promotes a decondensed chromatin environment, where enhancers and promoters can contact each other over longer distances, and where the resetting of enhancer-promoter interactions brought about by the fast decondensed chromatin motions is reduced, leading to more stable, long-lived enhancer-promoter relationships.

The role of dopamine D4 receptor in the induction of behavioral sensitization to amphetamine and accompanying biochemical and molecular adaptations.

Our studies examined the role of dopamine D4 receptors in the induction of behavioral sensitization to amphetamine (Amp) and accompanying neurochemical and molecular adaptive responses using a highly selective D4 antagonist, PNU-101387G. Behavioral sensitization to an acute challenge of Amp (2 mg/kg, s.c.) was observed in rats pretreated with five daily doses of Amp (2 mg/kg/d, s.c.) followed by 7-day withdrawal. Interestingly, coadministration of PNU-101387G with Amp during pretreatment completely blocked the sensitized response to an acute Amp challenge. The behavioral sensitization and its blockade by the D4 antagonist were observed in the absence of significant differences in cerebellar Amp levels among the various pretreatment groups. Accompanying behavioral sensitization were two postsynaptic neuroadaptive responses: reduction in the ability of Amp to induce c-fos gene expression in the infralimbic/ventral prelimbic cortex and NT/N mRNA in the accumbal shell. However, concurrent blockade of D4 receptors during Amp pretreatment prevented the refractoriness in c-fos and NT/N responsiveness to acute Amp. We observed also a presynaptic neuroplastic response associated with the behavioral sensitization: a significant augmentation in the ability of Amp to increase extracellular dopamine concentrations in the nucleus accumbens shell. As with the behavioral sensitization and associated postsynaptic adaptive responses, concurrent administration of PNU-101387G with Amp during pretreatment blocked the augmentation in Amp-induced dopamine release. Taken together, these data demonstrate that dopamine D4 receptors play an important role in the induction of behavioral sensitization to Amp and accompanying adaptations in pre- and postsynaptic neural systems associated with the mesolimbocortical dopamine projections.

Anxiolytic-like effects of PNU-101017, a partial agonist at the benzodiazepine receptor.

PNU-101017 is a chemically novel ligand at the benzodiazepine recognition site of cloned GABAA receptors. It was reported to potentiate GABA-mediated chloride current in cultured cells with a moderate intrinsic activity and a biphasic dose-response relationship. In this study, we confirmed that PNU-101017 has a partial agonist-like effect in the antagonism of metrazole-induced seizures in mice. It produced no sedation or ataxia, but did antagonize diazepam-induced motor deficit of mice in the rotarod test. PNU-101017 was weakly active in anti-conflict anxiolytic tests, but attenuated the plasma corticosteroid response to mild stress in rats. It also antagonized stress-induced elevation of cerebellar cGMP levels in mice. Like chlordiazepoxide, it increased drinking of saline solution in thirsty rats. PNU-101017 did not potentiate the CNS-depressant effects of ethanol, and produced no evidence of physical dependence when administered repeatedly. Agonists with low intrinsic activity at the benzodiazepine receptor, such as PNU-101017, should be further explored for therapeutic uses.