The DRD2 TaqI-B polymorphism and its relationship to smoking abstinence and withdrawal symptoms.

The dopamine receptor D2 (DRD2) gene has polymorphisms that have been linked to regulation of the dopamine system and to an increased prevalence of smoking. The present study examined the relationship of the DRD2 TaqI-A and -B polymorphisms with short-term clinical outcome (abstinence and withdrawal symptoms), collected from daily (14 pre-quit and 42 post-quit) diary data among smokers (n=116) treated with the nicotine patch plus either venlafaxine or placebo. The results showed that B1/B1 or B1/B2 smokers were slightly less likely to be abstinent on a given day than those homozygous for the TaqI-B2 allele. Significant DRD2 TaqI-B x time interactions were found for several of the withdrawal scales, indicating that those smokers with the B1/B1 or B1/B2 genotypes tended to report more symptoms over time compared to those with the B2/B2 genotype. No interactions or main effects were found for the DRD2 TaqI-A polymorphism. The findings demonstrate that smokers homozygous for the TaqI-B2 allele experience progressive improvement in self-reported withdrawal symptoms while smokers with the TaqI-B1 allele showing little change.

Healthcare providers' sun-protection promotion and at-risk clients' skin-cancer-prevention outcomes.

BACKGROUND: This study aims to determine whether healthcare providers' (HCPs') communication dealing with sun-protection (i.e., counseling) is associated with clients' skin-cancer-related prevention practices, detection self-efficacy, and knowledge. METHODS: Secondary analysis of two surveys of 1,469 randomly sampled farmers and soccer participants from southeast and coastal Georgia. RESULTS: Farmers and soccer participants who report ever having been counseled by a HCP about how to protect their skin from the sun report being more likely to wear sunscreen (P < 0.05), get clinical exams of their skin (P < 0.001), be certain that they can recognize unhealthy changes in their skin (P < 0.001), be certain that they know how to perform a skin exam (P < 0.001), and be knowledgeable about skin cancer prevention (P < 0.05 and P < 0.001, respectively); soccer participants are additionally more likely to wear protective headgear (P < 0.05) and perform monthly self-exams of their skin (P < 0.001). All analyses incorporated three control variables: participants' prior history of skin cancer, age, and non-HCP-derived skin-cancer awareness. CONCLUSIONS: Findings suggest that HCPs' counseling can positively shape skin-cancer-related prevention practices, detection self-efficacy, and knowledge. Additional research is needed on HCPs' actual communication about skin cancer and sun protection and its influence on client outcomes.

Women's health problems in soap operas: a content analysis.

A content analysis of women's health problems in network daytime serials is reported. Results indicate a preponderance of such problems as substance abuse, falls resulting in death or injury, comas or unconsciousness, mental health concerns (most notably represented by amnesia and multiple personality), concerns about AIDS, automobile accidents, being drugged, fainting, pregnancy problems, and violent acts resulting in harm. Such concerns as breast and cervical cancer and heart disease are less frequently portrayed.

Transient kinetics of substrate binding to Na+/K(+)-ATPase measured by fluorescence quenching.

This paper examines the transient kinetics of substrate binding to the Na+/K(+)-ATPase labelled with iodoacetamidofluorescein (IAF) using fluorescence quenching by trinitrophenyl-ATP (TNP-ATP). Earlier work (E.H. Hellen, P.R. Pratap, 1996, Fluorescence quenching of IAF-Na+/K(+)-ATPase via energy transfer to TNP-labelled nucleotide, Proceedings of the VIIIth International Conference on the Na+/K(+)-ATPase, in press) has shown that TNP-nucleotide binds to specific sites (from which unlabelled nucleotide can displace it) and nonspecific sites (from which unlabelled nucleotide cannot displace it). Under stopped-flow conditions, quenching of IAF-enzyme fluorescence was well described by a stretched exponential (F(t) = F infinity + delta F exp[-Bt alpha]). Physically, this function may be interpreted in terms of its inverse Laplace transform phi (k), which describes a distribution of rate-constants; alpha reflects the width of this distribution. As TNP-ATP concentration increased, alpha decreased, reflecting TNP-ATP binding to sites with higher energy barriers. alpha decreased by about the same amount with increasing [TNP-ATP] in the presence of saturating ATP, indicating that the distribution of rate-constants is largely associated with the nonspecific binding sites. However, alpha was significantly less than 1 for ATP-induced fluorescence recovery in the presence of TNP-ATP, indicating that rate-constants associated with specific binding site are also distributed. The distribution of rate-constants for binding to the specific site indicates a distribution in the energy of the transition state for substrate binding. These results suggest that the specific binding site (in either the empty or the full state) may exist in a series of conformations separated by small energy barriers. However, the energy barriers for binding associated with these conformations are significantly distributed.

Kinetics of conformational changes associated with potassium binding to and release from Na+/K(+)-ATPase.

The Na+/K(+)-ATPase functions in cells to couple energy from the hydrolysis of ATP to the transport Na+ out and K+ in. The fluorescent probe IAF (iodoacetamidofluorescein) covalently binds to this enzyme, reporting conformational changes without inhibiting enzyme activity. This paper describes experiments using dog kidney enzyme labeled with IAF to examine kinetics of conformational changes resulting from added Na+ and K+, measured in terms of steady-state and stopped-flow fluorescence changes. Kinetics of these fluorescence changes were examined as a function of temperature from two initial conditions: (a) enzyme in the high-fluorescence form (E(high)) was rapidly mixed with varying [K+]; and (b) enzyme in the low-fluorescence form (E(low)) was rapidly mixed with varying [ATP]. These experiments showed: (1) The rate constant for the fluorescence change from E(high) to E(low) was much larger than that for the opposite transition, E(low) to E(high); (2) the apparent free energy of activation (Ea(app)) for the two transitions were different (as estimated from Arrhenius plots); (3) under steady-state conditions, IAF fluorescence did not change when ATP was added to E(low)(K+) in the absence of Na+; (4) the apparent free energy of activation was independent of [K+] for the E(high) to E(low) transition (at 16.4 kcal/mol) but increased with [ATP] for the E(low) to E(high) transition; (5) Ea(app) for the E(low) to E(high) transition with 1 mM ATP was approximately the same as that in the absence of ATP (34 kcal/mol). These results can be interpreted as: (i) in the transition from E(low) to E(high), IAF reported a conformational change that occurred after K+ release to the intracellular side and which is involved in Na+ binding; (ii) Ea(app) increased with [ATP], while increasing the entropy of the transition state. Thus, ATP appeared to destabilize the enzyme during the transition from E(low) to E(high).

Rapid kinetic analyses of the Na+/K(+)-ATPase distinguish among different criteria for conformational change.

The Na+/K(+)-ATPase couples the hydrolysis of ATP to the transport of Na+ and K+ via a phosphorylated intermediate and conformational changes. In order to identify these conformational changes, we have probed the sequence of steps from EP(3Na+ in) to EP + 3Na+ out with three fluorescent probes (IAF: 5-iodoacetamidofluorescein; BIPM: N-[p-(2-benzimidazolyl)phenyl]maleimide; and RH421) and the sensitivity of their fluorescence change to oligomycin and divalent cations (Ca2+ and Mn2+). The magnitude (% delta F) and rate constant (k(obs)) of ATP-induced fluorescence changes were measured on a fluorescence stopped-flow apparatus, and yielded the following results. (a) With RH421, k(obs) and % delta F varied with [Na+] (maximal k(obs) = 100 s-1, K1/2 = 6 mM; % delta Fmax = 6%, K1/2 = 1 mM); these values are comparable to those previously reported using IAF-labeled enzyme (Pratap, P.R., Robinson, J.D. and Steinberg, M.I. (1991) Biochim. Biophys. Acta 1069, 288-298). (b) With BIPM-labeled enzyme k(obs) did not vary with [Na+] over the range tested, and was twice as high as the maximum k(obs) for RH421. (c) Treatment with oligomycin reduced k(obs) for all three probes to about the same level (approximately 1-2 s-1) while % delta Fmax was largely unaffected. (d) Replacing Mg2+ with Ca2+ had similar effects to treatment with oligomycin. (e) RH421 fluorescence change was completely abolished in the presence of oligomycin and Ca2+. (f) Replacing Mg2+ with Mn2+ decreased IAF fluorescence, i.e., put the enzyme in an E2-like form, reduced k(obs), and rendered oligomycin less effective in reducing k(obs). From these results, we conclude: (a) the release of the second/third Na+ is the rate-limiting step for the conformational change measured by IAF and charge transfer measured with RH421; (b) BIPM indicates an earlier step, either the deocclusion of Na+ and/or the release of the first Na+; (c) oligomycin blocks Na+ deocclusion, and this step is sensitive to the divalent cation used to activate enzyme phosphorylation; and (d) Ca2+ slows the step reported by IAF as well. These experiments indicate that a simple model with two conformations (E1 and E2) is insufficient to explain transient kinetic data.

Improved mammography with a reduced radiation dose.

A 2-cm-high focused air-interspaced grid with a 7:1 ratio and eight lamellae per inch was devised. The grid is arc-shaped and pivots on its focal point, the anode-cathode axis. Milliamperage was 35% less for breast phantom radiographs obtained with the new grid versus those obtained with a conventional 5:1 mammographic grid. In a comparison study of five mammograms, each evaluated for five parameters, the images obtained with the new grid were judged to be of equal or better quality than those obtained with a conventional grid in 17 of 20 comparisons.

Effects of choline on Na(+)- and K(+)-interactions with the Na+/K(+)-ATPase.

Choline chloride, 100 mM, stimulates Na+/K(+)-ATPase activity of a purified dog kidney enzyme preparation when Na+ is suboptimal (9 mM Na+ and 10 mM K+) and inhibits when K+ is suboptimal (90 mM Na+ and 1 mM K+), but has a negligible effect at optimal concentrations of both (90 mM Na+ and 10 mM K+). Stimulation occurs at low Na+ to K+ ratios, but not at those same ratios when the actual Na+ concentration is high (90 mM). Stimulation decreases or disappears when incubation pH or temperature is increased or when Li+ is substituted for K+ or Rb+. Choline+ also reduces the Km for MgATP at the low ratio of Na+ to K+ but not at the optimal ratio. In the absence of K+, however, choline+ does not stimulate at low Na+ concentrations: either in the Na(+)-ATPase reaction or in the E1 to E2P conformational transition. Together, these observations indicate that choline+ accelerates the rate-limiting step in the Na+/K(+)-ATPase reaction cycle, K(+)-deocclusion; consequently, optimal Na+ concentrations reflect Na+ accelerating that step also. Thus, the observed K0.5 for Na+ includes high-affinity activation of enzyme phosphorylation and low-affinity acceleration of K(+)-deocclusion. Inhibition of Na+/K(+)-ATPase and K(+)-nitrophenylphosphatase reactions by choline+ increases as the K(+)-concentration is decreased; the competition between choline+ and K+ may represent a similar antagonism between conformations selected by choline+ and by K+.

The reaction sequence of the Na+/K(+)-ATPase: rapid kinetic measurements distinguish between alternative schemes.

Conformational changes between E1 and E2 enzyme forms of a dog kidney Na+/K(+)-ATPase preparation labeled with 5-iodoacetamidofluorescein were followed with a stopped-flow fluorimeter, in terms of the rate constant, kobs, and the steady-state magnitude, % delta F of fluorescence change. On rapid mixing of enzyme plus Mg2+ plus Na+ with saturating (0.5 mM) ATP in the absence of K+, kobs varied with Na+ concentration in the range 0-155 mM, with a K1/2 of 10 mM, while % delta F was relatively insensitive to Na+, with a K1/2 of 0.5 mM. Oligomycin reduced kobs by 98-99% for Na+ greater than or equal to 10 mM, but only by 50% for Na+ = 1 mM; % delta F was reduced at most by 20%. At 155 mM Na+, both kobs and % delta F changed if K+ was present with the enzyme. kobs decreased by 50% when K+ was increased from 0 to 0.2 mM, but increased when K+ was varied in the range 0.2-5 mM. K+ increased % delta F by a factor of 3 with a K1/2 of 0.3-0.5 mM as measured in both stopped-flow and steady-state experiments. These data are considered in terms of the derived presteady-state equations for two alternate schemes for the enzyme, with the E1P to E2P conformational change either preceding (Albers-Post) or following (Nørby-Yoda-Skou) Na+ transport and release. The analysis indicates that: (i) Na+ must be released before the conformational transition, from an E1 form; (ii) the step in which the second and/or third Na+ is released is rate-limiting, but this release is accelerated by Na+; and (iii) the release is also accelerated by K+ acting with low affinity (possibly at extracellular sites).

Na+/K(+)-ATPase: modes of inhibition by Mg2+.

Adding 15 mM free Mg2+ decreased Vmax of the Na+/K(+)-ATPase reaction. Mg2+ also decreased the K0.5 for K+ activation, as a mixed inhibitor, but the increased inhibition at higher K+ concentrations diminished as the Na+ concentration was raised. Inhibition was greater with Rb+ but less with Li+ when these cations substituted for K+ at pH 7.5, while at pH 8.5 inhibition was generally less and essentially the same with all three cations: implying an association between inhibition and ion occlusion. On the other hand, Mg2+ increased the K0.5 for Na(+)-activation of the Na+/K(+)-ATPase and Na(+)-ATPase reactions, as a mixed inhibitor. Changing incubation pH or temperature, or adding dimethylsulfoxide affected inhibition by Mg2+ and K0.5 for Na+ diversely. Presteady-state kinetic studies on enzyme phosphorylation, however, showed competition between Mg2+ and Na+. In the K(+)-phosphatase reaction catalyzed by this enzyme Mg2+ was a (near) competitor toward K+. Adding Na+ with K+ inhibited phosphatase activity, but under these conditions 15 mM Mg2+ stimulated rather than inhibited; still higher Mg2+ concentrations then inhibited with K+ plus Na+. Similar stimulation and inhibition occurred when Mn2+ was substituted for Mg2+, although the concentrations required were an order of magnitude less. In all these experiments no ionic substitutions were made to maintain ionic strength, since alternative cations, such as choline, produced various specific effects themselves. Kinetic analyses, in terms of product inhibition by Mg2+, require Mg2+ release at multiple steps. The data are accommodated by a scheme for the Na+/K(+)-ATPase with three alternative points for release: before MgATP binding, before K+ release and before Na+ binding. The latter alternatives necessitate two Mg2+ ions bound simultaneously to the enzyme, presumably to divalent cation-sites associated with the phosphate and the nucleotide domains of the active site.

Interventional magnetic resonance imaging in the head and neck.

Interventional MRI is clearly in its early stages of development. While the value of MR-guided aspiration cytology and MR evaluation of deep electrode implantation in the brain has already been confirmed with human clinical studies, the future of MR-guided interstitial laser therapy remains to be proven. Despite this, as we look ahead into the 1990s and the millennium, it is possible to imagine dedicated MR laser therapy units for combined radiological and surgical outpatient approaches in what may become the operating rooms of the 21st century.

Interstitial laser phototherapy assisted by magnetic resonance imaging: a new technique for monitoring laser-tissue interaction.

The rapid technological advances of magnetic resonance imaging, laser fiberoptics, and compatible probes may allow treatment of deep and sometimes surgically unreachable tumors of the head and neck with minimal morbidity through interstitial laser phototherapy. In this study, a new application of magnetic resonance imaging was developed to monitor and quantify laser-induced tissue damages. Pig skin was exposed to increased levels of argon laser (514.5 nm) at energy densities between 62.5 and 375 J/cm2 as determined by an accurate and reproducible method of dosimetry. Thermal profiles were recorded using an infrared sensor and T1- and T2-weighted magnetic resonance images were taken; afterward, biopsies were performed to quantitate the level of tissue damage. Our results demonstrate that above a certain threshold of laser energy, the magnetic resonance imaging findings are temperature dependent. Appropriate development of a scale matching laser energies, temperature profiles, T1- and T2-weighted magnetic resonance images, and histological quantitation of tissue destruction will allow us to optimize the three-dimensional control and monitoring of laser-tissue interactions.

Differential effects of substrates on three transport modes of the Na+/K(+)-ATPase.

With a purified Na+/K(+)-ATPase preparation reconstituted into phospholipid vesicles, Na+/K+, Na+/Na+, and uncoupled Na+ transport were studied using three nucleotides and five substrates of the K(+)-phosphatase reaction that this enzyme also catalyzes. For Na+/K+ exchange, CTP was half as effective as ATP and GTP one-twentieth; of the phosphatase substrates only carbamyl phosphate and 3-O-methylfluorescein phosphate produced significant transport and at merely 1% of the rate with ATP. For Na+/Na+ exchange, comparable rates of transport were produced by ATP, CTP, carbamyl phosphate and acetyl phosphate, although the actual rate of transport with ATP was only 2.4% of that for Na+/K+ exchange; slower rates occurred with GTP (69%), 3-O-methylfluorescein phosphate (51%), and nitrophenyl phosphate (33%). Only umbelliferone phosphate was ineffective. For uncoupled Na+ transport results similar to those for Na+/Na+ exchange were obtained, but the actual rate of transport was still slower, 1.4% of that for Na+/K+ exchange. Thus, not only nucleotides but a variety of phosphatase substrates (which are phosphoric acid mixed anhydrides) can phosphorylate the enzyme at the high-affinity substrate site to form the E1P intermediate of the reaction sequence. Oligomycin inhibited Na+/K+ exchange with ATP by half, but with carbamyl phosphate not at all; with CTP the inhibition was intermediate, one-fourth. By contrast, oligomycin inhibited Na+/Na+ exchange by one-fifth with all three substrates. A quantitative, steady-state kinetic model accounts for the relative magnitudes of Na+/K+ and Na+/Na+ exchanges with ATP, CTP, and carbamyl phosphate as substrates, as well as the extents of inhibition by oligomycin. The model requires that even when Na+ substitutes for K+ a slow step in the reaction sequence is the E2 to E1 conformational transition.

The biocompatibility of compressed collagen foam plugs.

The tissue reaction to reexpanded, purified bovine collagen sponge placed percutaneously into the lung, pleural space, liver, kidney, and muscle was studied in dogs and rabbits. In addition, the biocompatibility and radiopacity of tantalum-treated collagen foam plugs was examined. No adverse effects were found. We believe that collagen plugs may be of use in occluding needle tracts from biopsy sites to prevent complications such as bleeding or pneumothoraces.

Volume growth rate of acoustic neuromas on MRI post-stereotactic radiosurgery.

Of the approximately 160 acoustic neuroma patients treated with stereotactic radiosurgery in the world up to 1987, 8 patients at UCLA Medical Center have had two or more magnetic resonance scans at least one year apart available for study (all 8 patients were treated with stereotactic radiosurgery for acoustic neuromas by the Department of Neurosurgery at the Karolinska Hospital, Stockholm, Sweden). The followup time after radiosurgery ranged from 4 to 8 years. The volume doubling rate post-stereotactic radiosurgery was calculated to be slow (763 to 888 days) in two patients, virtually arrested in five patients (doubling times larger than 2500 days) and negative (-563 days) in one patient indicating a shrinking tumor. Due to the limited sample size no radiological finding or clinical data correlated with the volume doubling times. A control patient that had no treatment for her tumor had a doubling time of 217 days for comparison.

[The reaction mechanism of Na, K-ATPase]. / Mekhanizm reaktsii Na, K-ATFazy.

To help characterize the Na,K-ATPase active site with enzyme incorporated into phospholipid vesicles, the activities with alternative substrates were compared, 22Na/Na-transport was equivalent with ATP, CTP, carbamylphosphate and acetylphosphate, but slower with CTP, 3-O-methylfluoresceinphosphate (3-O-MFP), nitrophenylphosphate and umbelliferonephosphate. It indicates a slower rate of formation of phosphorylating enzyme complex in conformation position of E1 (E1P) when the second group of substrates is bound with enzyme active center. 22Na/K-transport was half as effective with CTP as with ATP and was far slower with the other substrates. It indicates a more stringent selectivity at the low-affinity site of enzyme in conformation E2 that accelerates the slow step of this transport mode. Although enzyme modification with fluoresceinisothiocyanate blocks the high-affinity site to ATP, the K-phosphatase reaction catalyzed by E2 is retained, even with a substrate, 3-O-MFP, that binds to the adenine pocket. Dimethylsulfoxide inhibits hydrolysis of the nucleotides and of the carboxylic phosphate substrates of the K-phosphatase reaction, but stimulates hydrolysis of the phenolic phosphate substrates (nitrophenylphosphate and umbelliferone phosphate) which normally are hydrolyzed more slowly than the other substrates. On the basis of these data the authors propose the model of Na,K-ATPase active center.

Modification of ligand binding to the Na+/K+-activated ATPase.

Interactions between the ligands Mg2+, K+, and substrate and the Na+/K+-activated ATPase were examined in terms of a rapid-equilibrium, random-order, terreactant kinetic scheme for the K+-nitrophenyl phosphatase reaction that is catalyzed by this enzyme. At 37 degrees C and pH 7.5 the derived values for the dissociation constants from the free enzyme were 0.2, 0.08, and 1.4 mM for Mg2+, K+, and substrate, respectively. For Mg2+ interactions, the presence of 20% (v/v) dimethyl sulfoxide (Me2SO) increased the calculated affinity 25-fold; higher concentrations increased affinity still further. Neither reducing the temperature to 20 degrees C nor altering the pH from 6.5 to 8.3 appreciably changed the affinity for Mg2+ in the absence or presence of Me2SO. The Mg2+ sites are thus characterized by an absence of functional groups ionizable in the pH range 6.5-8.3, with binding driven by entropy changes, and with Me2SO, probably through solvation effects on the protein, increasing affinity for Mg2+ close to that for Ca2+ and Mn2+. By contrast, for K+ interactions, the presence of 20% Me2SO increased the calculated affinity only by half; moreover, reducing the temperature to 20 degrees C and the pH to 6.5 both increased affinity and diminished the response to Me2SO. The K+ sites are thus characterized by a marked sensitivity to pH and temperature, presumably through alterations in enzyme conformational equilibria that in turn are modifiable by Me2SO. Inhibition by higher concentrations of Mg2+, which varies inversely with the K+ concentration, was decreased by Me2SO. Finally, for substrate interactions, the presence of 20% Me2SO increased the calculated affinity 4-fold, and, as for Mg2+-binding, neither reducing the temperature nor varying the pH over the range 6.5-8.3 appreciably altered the affinity in the absence or presence of Me2SO. Thus, the substrate sites, like the Mg2+ sites, are characterized by an absence of functional groups ionizable in this range, with binding driven by entropy changes, and with Me2SO increasing affinity for substrate, in this case probably through favoring the partitioning of substrate from the medium into the hydrophobic active site.