Publications

To study conformational transitions at the muscle nicotinic acetylcholine (ACh) receptor (nAChR), a rhodamine fluorophore was tethered to a Cys side chain introduced at the beta 19' position in the M2 region of the nAChR expressed in Xenopus oocytes. This procedure led to only minor changes in receptor function. During agonist application, fluorescence increased by (Delta F/F) approximately 10%, and the emission peak shifted to lower wavelengths, indicating a more hydrophobic environment for the fluorophore. The dose-response relations for Delta F agreed well with those for epibatidine-induced currents, but were shifted approximately 100-fold to the left of those for ACh-induced currents. Because (i) epibatidine binds more tightly to the alpha gamma-binding site than to the alpha delta site and (ii) ACh binds with reverse-site selectivity, these data suggest that Delta F monitors an event linked to binding specifically at the alpha delta-subunit interface. In experiments with flash-applied agonists, the earliest detectable Delta F occurs within milliseconds, i.e., during activation. At low [ACh] (< or = 10 microM), a phase of Delta F occurs with the same time constant as desensitization, presumably monitoring an increased population of agonist-bound receptors. However, recovery from Delta F is complete before the slowest phase of recovery from desensitization (time constant approximately 250 s), showing that one or more desensitized states have fluorescence like that of the resting channel. That conformational transitions at the alpha delta-binding site are not tightly coupled to channel activation suggests that sequential rather than fully concerted transitions occur during receptor gating. Thus, time-resolved fluorescence changes provide a powerful probe of nAChR conformational changes.

BACKGROUND

A longstanding but controversial hypothesis is that individuals who exhibit frequent, large increases in blood pressure (BP) during psychological stress are at risk for developing essential hypertension. We tested whether BP changes during psychological stress predict incident hypertension in young adults.

METHODS AND RESULTS

We used survival analysis to predict hypertensive status during 13 years of follow-up in a sample of >4100 normotensive black and white men and women (age at entry, 18 to 30 years) enrolled in the CARDIA study. BP responses to 3 psychological challenges--cold pressor, star tracing, and video game tasks--were measured. Hypertensive status was defined as use of antihypertensive medication or measured BP > or =140/90 mm Hg. After adjustment for race, gender, covariates (education, body mass index, age, and resting pressure), and their significant interactions, the larger the BP responses were to each of the 3 tasks, the earlier hypertension occurred (P<0.0001 to <0.01). The systolic BP effect for the cold pressor task was apparent for women and for whites in race- and gender-specific models, whereas the diastolic BP effect for the video game was apparent for men.

CONCLUSIONS

Young adults who show a large BP response to psychological stress may be at risk for hypertension as they approach midlife.