Biochemistry has led the way in reducing what once were considered irreducible biological phenomena to fundamental physico-chemical principles. Keys to this accomplishment were the ability to trace the transformations of defined chemical substrates in cellular extracts or intact cells and the reconstitution of biochemical processes with pure components. For neuroscience to follow this reductionist lead, analogous experimental strategies need to be developed: an ability to trace the transformations of defined neuronal activity patterns in explanted neural tissues or intact nervous systems and a capacity to reconstitute neurally encoded information and the behaviors it guides with synthetic neural signals.The initial hurdle to both of these strategies lies in the difficulty of feeding artificial neural signals into functionally circumscribed but anatomically dispersed populations of neurons. An organic solution to this problem is to harness proteins mediating neuronal excitation as conduits for artificial stimuli and to restrict the expression of these transducers genetically to a predetermined group of target neurons (1, 2). If, for example, a set of generalist neurons could be programmed to express signal transduction machinery that normally is present only in specialized sensory cells, these neurons also might acquire the capacity to respond selectively to the adequate physical or chemical triggers. Not only could all members of a population of neurons then be addressed simultaneously, susceptibility to stimulation rather than the stimulus itself would be localized, and even diffusely broadcast stimuli could elicit precise, patterned responses (1, 2).Neurons offer two principal routes for the transduction of excitatory signals (3). Metabotropic signaling systems consist of heptahelical receptors that communicate with their effectors through heterotrimeric G proteins. The activation of some of these effectors is coupled to changes in membrane potential (3-7). Ionotropic signaling systems effect changes in membrane potential directly, via chemically or physically gated conductances (3). Previously, we have used components of a metabotropic system, the phototransduction cascade of the fruit fly (7), to sensitize vertebrate neurons to light (2). Expression of what was termed ''chARGe'' to commemorate the essential elements (arrestin-2, rhodopsin, and a G protein ␣-subunit) created a light-controlled source of depolarizing current whose activation sparked action potentials (2). We now introduce the use of ionotropic mechanisms for the selective chemical and optical stimulation of genetically designated populations of neurons. In addition to multiple trigger modes, these ionotropic systems possess numerous practical advantages over chARGe, including simplicity, fast kinetics, and broad tunability. Materials and MethodsHeterologous Expression of Ion Channels. Candidate ion channels were expressed under the control of the cytomegalovirus promoter in pCI-fluor, a derivative of the mammalian expression vector pCI-neo (Pr...

A general microscale protocol for the determination of absolute configurations of primary amino groups or secondary hydroxyl groups linked to a single stereogenic center is described. The chiral substrates are linked to the achiral trifunctional bidentate carrier molecule (3-aminopropylamino)acetic acid (1, H(2)NCH(2)CH(2)CH(2)NHCH(2)COOH) and the resultant conjugates are then complexed with dimeric zinc porphyrin host 2 giving rise to 1:1 host/guest sandwiched complexes. These complexes exhibit exciton-coupled bisignate CD spectra due to stereodifferentiation leading to preferred porphyrin helicity. Since the chiral sense of twist between the two porphyrins in the complex is dictated by the stereogenic center of the substrate, the sign of the couplet determines the absolute configuration at this center. The twist of the porphyrin tweezer in the complex can be predicted from the relative steric sizes of the groups flanking the stereogenic center, such that the bulkier group protrudes from the complex sandwich. In certain alpha-hydroxy esters and alpha-amino esters, electronic factors and hydrogen bonding govern the preferred conformation of the complex, and hence the CD spectra.

We recently reported a new method for the direct dehydrogenative C-H silylation of heteroaromatics utilizing Earth-abundant potassium tert-butoxide. Herein we report a systematic experimental and computational mechanistic investigation of this transformation. Our experimental results are consistent with a radical chain mechanism. A trialkylsilyl radical may be initially generated by homolytic cleavage of a weakened Si-H bond of a hypercoordinated silicon species as detected by IR, or by traces of oxygen which can generate a reactive peroxide by reaction with [KOt-Bu] as indicated by density functional theory (DFT) calculations. Radical clock and kinetic isotope experiments support a mechanism in which the C-Si bond is formed through silyl radical addition to the heterocycle followed by subsequent β-hydrogen scission. DFT calculations reveal a reasonable energy profile for a radical mechanism and support the experimentally observed regioselectivity. The silylation reaction is shown to be reversible, with an equilibrium favoring products due to the generation of H gas. In situ NMR experiments with deuterated substrates show that H is formed by a cross-dehydrogenative mechanism. The stereochemical course at the silicon center was investigated utilizing a H-labeled silolane probe; complete scrambling at the silicon center was observed, consistent with a number of possible radical intermediates or hypercoordinate silicates.

Sulfonamide antimicrobials are used in both human therapy and animal husbandry. Sulfonamides are not readily biodegradable and have been detected in surface water and in secondary wastewater effluents. The chemical oxidation of sulfonamides by an environmentally friendly oxidant, ferrate(VI) (Fe(VI)O4(2-), Fe(VI)), was conducted. The sulfonamides used in the oxidation studies were sulfisoxazole, sulfamethazine, sulfamethizole, sulfadimethoxine, and sulfamethoxazole. Kinetics of the reactions were determined as a function of pH (7.0-9.7) and temperature (15-45 degrees C) by a stopped-flow technique. The rate law for the oxidation of sulfonamides by Fe(VI) is first-order with respect to each reactant. The observed second-order rate constants decreased nonlinearly with an increase in pH and are possibly related to the protonation of Fe(VI) (HFeO4- <==> H+ + FeO4(2-); pK(a,HFeO4) = 7.23) and sulfonamides (SH <==> H+ + S-; pK(a,SH) = 5.0-7.4). The activation parameters of the reactions vary with pH due to temperature dependence on the protonation of Fe(VI) and sulfonamides. These results were used to obtain enthalpy of dissociation of sulfonamides. Stoichiometry and products of sulfamethoxazole (SMX) reactions with Fe(VI) were studied in detail using various analytical techniques to evaluate the effect of the oxidation process on the fate of sulfonamides in water. At a stoichiometric ratio of 4:1 (Fe(VI): SMX), complete removal of SMX was achieved. Analyses of oxidation products of the reaction as well as kinetic measurements of substructural models of SMX suggest that the attack of Fe(VI) occurs at the isoxazole moiety as well as at the aniline moiety with minimal preference. The results of the studies reported suggest that Fe(VI) has the potential to serve as a chemical oxidant for removing sulfonamides and converting them to relatively less toxic byproducts in water.