Hyperpolarization

The main function of background two-pore potassium (K(2P)) channels KCNK3/9/15 is to stabilize the cell membrane potential. We previously observed that membrane potential depolarization enhances the release of HIV-1 viruses. Because membrane polarization affects the biomembrane directly, here we examined the effects of KCNK3/9/15 on the budding of nonviral vesicles. We found that depolarization by knocking down endogenous KCNK3/9/15 promoted secretion of cell-derived vesicles. We further used Vpu (an antagonist of KCNK3) as a model for the in vivo study of depolarization-stimulated secretion. Vpu is a HIV-1-encoded, ion channel-like protein (viroporin) capable of enhancing virus release and depolarizing the cell membrane potential. We found that Vpu could also promote nonviral vesicle release, perhaps through a similar mechanism that Vpu utilizes to promote viral particle release. Notably, T cells expressing Vpu alone became pathologically low in intracellular K(+) and insensitive to extracellular K(+) or membrane potential stimulation. In contrast, heterologous expression of KCNK3 in T cells stabilized the cell potentials by maintaining intracellular K(+). We thus concluded that KCNK3/9/15 expression limits membrane depolarization and depolarization-induced secretion at least in part by maintaining intracellular K(+).

A set of new methacrylate monomers based on azobenzene/azomethine moieties coupled with an antipyrine heterocyclic fragment was designed. Their free radical polymerization provided six homo-/ copolymers with photoactive side chains. These novel materials were characterized with NMR, IR and UV-vis spectroscopies, as well as DSC and SEC. Their THF solutions demonstrated an efficient refractive nonlinear optical (NLO) response with an optical quality factor, FOM, of 10 2-2 Â 10 3 via self-action of single picosecond range laser pulses at a wavelength of 532 nm. Their efficiency enhances in the following set of chromophores: azomethine o azo o bis-azomethine. The polymers exhibit higher cubic hyperpolarizabilities against their corresponding initial molecules, while the fractional contribution of the photoactive group reduces. Incorporation of bis-structured units improves both the polymer properties and the NLO response efficiency of the materials. The fractional contribution of a photochromic group to the macromolecular refractive/absorptive NLO optical responses was analyzed for the first time.

The contribution of small-conductance (SKCa) and intermediate-conductance Ca2+-activated K+ (IKCa) channels to the generation of nitric oxide (NO) by Ca2+-mobilizing stimuli was investigated in human umbilical vein endothelial cells (HUVECs) by combining single-cell microfluorimetry with perforated patch-clamp recordings to monitor agonist-evoked NO synthesis, cytosolic Ca2+ transients, and membrane hyperpolarization in real time. ATP or histamine evoked reproducible elevations in NO synthesis and cytosolic Ca2+, as judged by 4-amino-5-methylamino-2′,7′-difluorofluorescein (DAF-FM) and fluo-3 fluorescence, respectively, that were tightly associated with membrane hyperpolarizations. Whereas evoked NO synthesis was unaffected by either tetraethylammonium (10 mmol/l) or BaCl2 (50 μmol/l) + ouabain (100 μmol/l), depleting intracellular Ca2+ stores by thapsigargin or removing external Ca2+ inhibited NO production, as did exposure to high (80 mmol/l) external KCl. Importantly, apamin and ...

The hypotensive and vasorelaxant effect of dioclein in resistance mesenteric arteries was studied in intact animals and isolated vessels, respectively.In intact animals, initial bolus administration of dioclein (2.5 mg kg−1) produced transient hypotension accompanied by an increase in heart rate. Subsequent doses of dioclein (5 and 10 mg kg−1) produced hypotensive responses with no significant change in heart rate. NG-nitro-L-arginine methyl ester (L-NAME) did not affect the hypotensive response.In endothelium-containing or -denuded vessels pre-contracted with phenylephrine, dioclein (5 and 10 mg kg−1 produced a concentration-dependent vasorelaxation (IC50=0.3±0.06 and 1.6±0.6 μM, respectively) which was not changed by 10 μM indomethacin. L-NAME (300 μM) produced a shift to the right.Dioclein was without effect on contraction of vessels induced by physiological salt solution (PSS) containing 50 mM KCl and the concentration dependence of dioclein's effect on phenylephrine induce...

Conventional magnetic resonance (MR) faces serious sensitivity limitations which can be overcome by hyperpolarization methods, but the most common method (dynamic nuclear polarization) is complex and expensive, and applications are limited by short spin lifetimes (typically seconds) of biologically relevant molecules. We use a recently developed method, SABRE-SHEATH, to directly hyperpolarize (15)N2 magnetization and long-lived (15)N2 singlet spin order, with signal decay time constants of 5.8 and 23 minutes, respectively. We find >10,000-fold enhancements generating detectable nuclear MR signals that last for over an hour. (15)N2-diazirines represent a class of particularly promising and versatile molecular tags, and can be incorporated into a wide range of biomolecules without significantly altering molecular function.

Upon hydrogenation from para-hydrogen (p-H(2)), hyperpolarization transfer toward a heteronucleus may be possible even if the two protons are chemically equivalent in the final product (but not magnetically equivalent), provided that J couplings with the heteronucleus exist. It is however shown (theoretically and experimentally) that this transfer effectively occurs if the spin system in the hydrogenated molecule is of the type AA'X (A and A' denoting the two protons originating from p-H(2) and X the heteronucleus) but does not occur for a spin system of the A(2)A(2)(')X type. A theory has been worked out for assessing the details of the X spectrum (multiplet patterns) in the case of ALTADENA and PASADENA experiments. Experimental verifications are provided.

Nuclear spin hyperpolarization makes a significant advance toward overcoming the sensitivity limitations of in vivo magnetic resonance imaging, particularly in the case of low-gamma nuclei. The sensitivity may be improved further by storing the hyperpolarization in slowly relaxing singlet populations of spin-1/2 pairs. Here, we report hyperpolarized 13C spin order transferred into and retrieved from singlet spin order using a small animal magnetic resonance imaging scanner. For spins in sites with very similar chemical shifts, singlet spin order is sustained in high magnetic field without requiring strong radiofrequency irradiation. The demonstration of robust singlet-to-magnetization conversion, and vice versa, on a small animal scanner, is promising for future in vivo and clinical deployments. Magn Reson Med, 2012. © 2012 Wiley Periodicals, Inc.

We show that optical excitation of radical triplet pair systems can produce a fourfold NMR signal enhancement in solution, without the need for microwave pumping. Development of optical hyperpolarization methods will significantly impact all NMR user groups by boosting sensitivity and reducing signal averaging times.