ABSTRACT
We present a hybrid single-molecule technique combining magnetic tweezers and Förster resonance energy transfer (FRET) measurements. Through applying external forces to a paramagnetic sphere, we induce conformational changes in DNA nanostructures, which are detected in two output channels simultaneously. First, by tracking a magnetic bead with high spatial and temporal resolution, we observe overall DNA length changes along the force axis. Second, the measured FRET efficiency between two fluorescent probes monitors local conformational changes. The synchronized orthogonal readout in different observation channels will facilitate deciphering the complex mechanisms of biomolecular machines.
Subject(s)
DNA/chemistry , Nanostructures/chemistry , Nanotechnology , Nucleic Acid Conformation , Fluorescence Resonance Energy Transfer , Fluorescent Dyes/chemistry , Magnetics , Optical TweezersABSTRACT
The endocannabinoid system (ECS) may either enhance or inhibit responses to aversive stimuli, possibly caused by its modulatory activity on diverse neurotransmitters. The aim of this work was to investigate the involvement of serotonin (5-HT) and catecholamines, as well as the role of glutamatergic and GABAergic cannabinoid type 1 (CB(1)) receptor, in responses to the antidepressant-like doses of the CB(1) receptor agonist Δ(9)-tetrahydrocannabinol (THC) and the antagonist rimonabant in the forced swim test (FST). Mice received acute injections of low doses of THC (0.1 or 0.5 mg/kg) or high dose of rimonabant (3 or 10 mg/kg) after treatment with the 5-HT synthesis inhibitor pCPA (100 mg/kg, 4 days), the 5-HT(1A) receptor antagonist WAY100635 (1 mg/kg, acute) or the non-selective blocker of catecholamine synthesis, AMPT (20 mg/kg, acute). THC and rimonabant were also tested in mutant mice lacking CB(1) receptor in specific forebrain neuronal subpopulations. Both THC and rimonabant induced antidepressant-like effects, quantified as immobility in the FST. However, only THC effects were reversed by pCPA or WAY100635. In contrast, only AMPT could attenuate the rimonabant effect. We also found decreased immobility in mice lacking the CB(1) receptor in glutamatergic cortical neurons, but not in forebrain GABAergic neurons, as compared with wild-type controls. The effect of THC persisted in mutant mice with CB(1) receptor inactivation in GABAergic neurons, whereas rimonabant effects were alleviated in these mutants. Thus, employing both pharmacological and genetic tools, we could show that the ECS regulates stress responses by influencing GABAergic, glutamatergic and monoaminergic transmission. The antidepressant-like action of THC depends on serotonergic neurotransmission, whereas rimonabant effects are mediated by CB(1) receptor on GABAergic neurons and by catecholamine signaling.
Subject(s)
Adaptation, Psychological/physiology , Neurons/physiology , Prosencephalon/physiology , Receptor, Cannabinoid, CB1/physiology , Serotonin/physiology , Stress, Psychological/drug therapy , Adaptation, Psychological/drug effects , Animals , Catecholamines/physiology , Dronabinol/pharmacology , Dronabinol/therapeutic use , GABAergic Neurons/drug effects , GABAergic Neurons/physiology , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Neurons/drug effects , Piperidines/pharmacology , Prosencephalon/drug effects , Pyrazoles/pharmacology , Receptor, Cannabinoid, CB1/agonists , Receptor, Cannabinoid, CB1/antagonists & inhibitors , Rimonabant , Stress, Psychological/psychologyABSTRACT
Allergy characterized by delayed hypersensitivity was induced in populations of mice by injection, respectively, of Staphylococcus aureus and of tubercle bacilli. Eliciting doses of staphylococcal antigen or of old tuberculin antigen were injected subcutaneously into the nape of the neck of samples of these mice. A challenge dose of S. aureus was injected into the site of the eliciting injection, and the subsequent survival curves of the infecting staphylococci were determined by plate counts. Survival of the challenge staphylococci was significantly reduced only when the eliciting antigen was homologous with the inducing antigen. Thus induction and elicitation were specific, but the local resistance was nonspecifically effective against staphylococci.
