Examining Sex Differences in Associations between Pain and Alcohol Use among College Student-Athletes.

Background: College student-athletes are a high-risk population for both pain and alcohol use. Although a growing literature indicates that pain motivates alcohol consumption, no studies have tested associations between pain and alcohol in college student-athletes. Methods and Results: Among National Collegiate Athletic Association Division I student-athletes at a large public university (N = 65; 48% Female), nearly all student-athletes (97%) reported alcohol use and 65% reported pain due to an athletic injury. Pain intensity was positively associated with greater motivation to reduce drinking prior to receiving a brief alcohol intervention. Among females, pain intensity was associated with lower self-reported alcohol consumption. Among male athletes, pain-related interference was associated with greater alcohol-related problems and lower perceived ability to deal with alcohol-related situations. Conclusion: Results provide initial evidence that pain may be important to consider when addressing alcohol use among student-athletes.

Comparison of Pregaming Alcohol Use and Consequences by Season Status and Sex in College Student Athletes.

As student athletes exhibit unique alcohol use patterns based on being in- versus out-of-season and biological sex, we aimed to explore student athlete (N = 442) alcohol use, pregaming behaviors, and associated negative outcomes. Results suggest being out-of-season and male are positively associated with negative alcohol-related consequences, and male athletes report greater numbers of pregame specific alcohol-related consequences than female athletes (p < .05). Female athletes indicated significantly higher estimated blood alcohol concentrations than male athletes on pregaming nights. No differences emerged between in- and out-of-season athletes on pregame consequences. Results suggest that further emphasis on the role season status and sex has on pregaming behaviors and experiencing negative outcomes may be an important next step toward enhancing prevention and intervention approaches.

The Influence of Exercise Intensity on Postexercise Baroreflex Sensitivity.

PURPOSE: The purpose of this study was to investigate the influence of exercise intensity on postexercise supine and tilt baroreflex sensitivity (BRS). METHOD: Nine healthy, active men performed 2 conditions of interval cycling of 40% maximal work rate (WRmax) and 75% WRmax of matched work done and a control condition of no exercise in a counterbalanced order. BRS outcome measures were determined at preexercise and postexercise up to +24 hr in supine and tilt positions. R-R interval and blood pressure data were collected over consecutive 10-min periods and were analyzed by Fast Fourier transformation analysis. RESULTS: A fully repeated analysis of variance revealed a significant interaction (p < .05) between time and condition in supine for BRSαLF, F(3, 134) = 5.19, p < .05, ES = .39, and BRSTFTG, F(3, 134) = 5.65, p < .05, ES = .41, and in tilt for BRSUpUp, F(3, 134) = 3.54, p < .05, ES = .31, BRSDownDown, F(3, 134) = 5.94, p < .05, ES = .43, BRSαLF, F(4, 134) = 6.23, p < .05, ES = .44, and BRSTFTG, F(4, 134) = 9.22, p < .05, ES = .54. There were significant differences (p < .05) between condition comparisons at +15 min and between control and 75% WRmax and between the 40% WRmax and 75% WRmax conditions at +60 min. At +15 min, BRS was lower in the 75% WRmax condition compared with the 40% WRmax condition and the control condition, and it was lower in the 40% WRmax condition than in the control condition. CONCLUSION: The findings demonstrate an intensity-dependent relationship in the BRS response following exercise.

Chronic treatment with a selective inhibitor of casein kinase I delta/epsilon yields cumulative phase delays in circadian rhythms.

INTRODUCTION: Casein kinase I epsilon/delta phosphorylates certain clock-related proteins as part of a complex arrangement of transcriptional/translational feedback loops that comprise the circadian oscillator in mammals. Pharmacologic inhibition leads to a delay of the oscillations with the magnitude of this effect dependent upon the timing of drug administration. OBJECTIVE: Earlier studies by our lab described the actions of a selective CKI epsilon/delta inhibitor, PF-670462, on circadian behavior following acute dosing; the present work extended these studies to chronic once-daily treatment. METHODS: Gross motor activity was used to estimate the circadian rhythms of rats maintained under a 12 L:12 D cycle. PF-670462, 10 or 30 mg/kg/day s.c., was administered once daily for 20 days either at ZT6 or ZT11 (i.e., 6 or 11 h after light onset). RESULTS: Chronic administration of PF-670462, performed at a fixed time of day, produced delays in the activity onsets of rats that cumulated with the duration of dosing. Dosing at ZT11 yielded more robust delays than dosing at ZT6 in keeping with earlier phase-response analyses with this agent. CONCLUSIONS: The magnitude of the shifts in activity onsets achieved with chronic dosing of PF-670462 appears to be a function of the dose and the previously established phase relationship. Its cumulative effect further suggests that the pharmacodynamic t (1/2) of the drug greatly exceeds its pharmacokinetic one. Most importantly, these changes in circadian behavior occurred in the presence of a fixed L:D cycle, confirming the drug to be a robust modulator of circadian phase in the presence of the natural zeitgeber.

Influence of the duration of a treadmill walking bout on heart rate variability at rest in physically active women.

BACKGROUND: Heart rate variability (HRV) has been promoted as a noninvasive method of evaluating autonomic influence on cardiac rhythm. Although female subjects predominate in the walking studies, no study to date has examined the influence of the duration of a moderate intensity walking physical activity bout on HRV in this population. METHODS: Twelve healthy physically active middle-aged women undertook 2 conditions; 20min (W20) and 60min (W60) bouts of walking on a treadmill. Resting HRV measures were obtained before (-1 h), and 1 h and 24 h after the walking bouts. RESULTS: Mean NN interval (ie, normal-to-normal intervals between adjacent QRS complexes) was significantly lower (P = .017) at +1 h in W60 (832, 686-979ms) compared with W20 (889, 732-1046ms). A borderline main effect for time was observed for both the SDNN intervals in W60 (P = .056), and for low frequency (LF(abs)) power in W60 (P = .047), with post hoc tests revealing a significant increase between -1 h (51, 33-69 ms and 847, 461-1556 ms(2)) and +1 h (65, 34-97ms and 1316, 569-3042 ms(2)) for SDNN and LF(abs) power, respectively, but no increase at +24h compared with -1 h. CONCLUSIONS: It appears that a walking bout of 60 min duration does alter cardiac autonomic influence in healthy active women, and this alteration is not evident after 20 min of walking. Given the rather subtle effect, further studies with larger sample sizes are required to explore the nature of the changes in cardiac autonomic influence following a prolonged bout of walking.

Inhibition of casein kinase I epsilon/delta produces phase shifts in the circadian rhythms of Cynomolgus monkeys.

INTRODUCTION: Circadian rhythms in mammals depend upon the cyclic oscillations of transcriptional/translational feedback loops in pacemaker cells of the suprachiasmatic nucleus. The rise and fall of clock-related proteins is a function of synthesis and degradation, the latter involving phosphorylation by casein kinase Iepsilon and delta. OBJECTIVE: Earlier studies by our lab described the actions of a selective CKIepsilon/delta inhibitor, PF-670462, on circadian behavior in rats; the present work extended these studies to a diurnal species, Cynomolgus monkeys. MATERIALS AND METHODS: General cage activity was used to estimate the circadian rhythms of eight telemeterized monkeys under baseline conditions and following s.c. doses of PF-670462. RESULTS AND DISCUSSION: Consolidated bouts of activity were noted during periods of light with a repeating period length of roughly 24 h based on their onset. Reassessment in constant dim light (42 vs. 450 lx) again yielded period lengths of 24 h, in this instance revealing the animals' endogenous rhythm. PF-670462 (10-100 mg/kg s.c.) produced a dose-dependent phase delay in much the same manner as that observed previously in rats. Dosing occurred 1.5 h prior to lights-off, roughly coincident with peaking levels of PER protein, a primary substrate of CKIepsilon/delta. CONCLUSIONS: These findings suggest that the time of dosing, when held constant in both the monkey and rat studies, produced nearly identical effects despite the subjects' diurnal or nocturnal preference. Importantly, these changes in rhythm occurred in the presence of light, revealing the drug as a powerful zeitgeber in a non-human primate and, by extension, in man.

An inhibitor of casein kinase I epsilon induces phase delays in circadian rhythms under free-running and entrained conditions.

Casein kinase Iepsilon (CKIepsilon) is an essential component of the biological clock, phosphorylating PER proteins, and in doing so regulating their turnover and nuclear entry in oscillator cells of the suprachiasmatic nucleus (SCN). Although hereditary decreases in PER phosphorylation have been well characterized, little is known about the consequences of acute enzyme inhibition by pharmacological means. A novel reagent, 4-[3-cyclohexyl-5-(4-fluoro-phenyl)-3H-imidazol-4-yl]-pyrimidin-2-ylamine (PF-670462), proved to be both a potent (IC(50) = 7.7 +/- 2.2 nM) and selective (>30-fold with respect to 42 additional kinases) inhibitor of CKIepsilon in isolated enzyme preparations; in transfected whole cell assays, it caused a concentration-related redistribution of nuclear versus cytosolic PER. When tested in free-running animals, 50 mg/kg s.c. PF-670462 produced robust phase delays when dosed at circadian time (CT)9 (-1.97 +/- 0.17 h). Entrained rats dosed in normal light-dark (LD) and then released to constant darkness also experienced phase delays that were dose- and time of dosing-dependent. PF-670462 yielded only phase delays across the circadian cycle with the most sensitive time at CT12 when PER levels are near their peak in the SCN. Most importantly, these drug-induced phase delays persisted in animals entrained and maintained in LD throughout the entire experiment; re-entrainment to the prevailing LD required days in contrast to the rapid elimination of the drug (t(1/2) = 0.46 +/- 0.04 h). Together, these results suggest that inhibition of CKIepsilon yields a perturbation of oscillator function that forestalls light as a zeitgeber, and they demonstrate that pharmacological tools such as PF-670462 may yield valuable insight into clock function.

Fluoxetine modulates the circadian biological clock via phase advances of suprachiasmatic nucleus neuronal firing.

BACKGROUND: The documented ability of serotonin (5-HT) to directly modulate circadian rhythms prompted interest in a similar role for therapeutic agents that readily enhance 5-HT neurotransmission, namely the selective serotonin reuptake inhibitors (SSRIs). METHODS: Extracellular recordings of unit firing of suprachiasmatic nucleus (SCN) neurons maintained in slice culture enabled determinations of circadian rhythmicity. Shifts in the peak of activity were determined during the next circadian cycle following drug exposure. RESULTS: Fluoxetine (10 microm, 60 minutes incubation) produced robust phase advances only in the presence of L-tryptophan (.5 microm), added to maintain serotonergic tone. CONCLUSIONS: Actions of SSRIs at the level of the circadian biological clock add to the list of pharmacological effects for this drug class and encourage speculation as to their importance clinically.

Circadian rhythm phenotype of 5-HT7 receptor knockout mice: 5-HT and 8-OH-DPAT-induced phase advances of SCN neuronal firing.

In vitro neuronal recordings in the SCN have clearly documented shifts in the peak of unit activity following the application of serotonergic agents, and yet selectivity issues with these very tools have limited progress in establishing the precise receptor mechanisms. As an alternative strategy, mice were bred (C57BL/6J) lacking 1 serotonin receptor, the 5-HT(7), to serve as a null background for this subtype; earlier work had documented the involvement of 5-HT(7) receptors in the phase advances elicited by 8-OH-DPAT, a mixed 5-HT(1A/7) agonist, in SCN slices prepared from rat donors. Single-unit recordings in sequential electrode passes revealed peaks of activity that occurred at nearly the same time in the knockout (KO; ZT4.2 +/- 0.6) and wild-type animals (WT; ZT4.3 +/- 0.1), where ZT0 marks the beginning of the light phase in a 12:12 LD cycle. Bath application of 8-OH-DPAT produced a phase advance in neuronal firing (2.1 +/- 0.5 h) when applied 1 circadian cycle earlier at ZT6 (10 microM, 10 min), but surprisingly, the mean phase advance in slices prepared from KO mice (2.3 +/- 0.1 h) was no different. Coapplication of 8-OH-DPAT with WAY-100,635 (10 microM), a highly selective 5-HT(1A) antagonist, significantly reduced the phase advance, both in experiments with WT and KO mice, suggesting the greater importance of this serotonin sub-type independent of genetic modification. 5-HT itself (0.5 +/-M, 10 min) at ZT6 also yielded phase advances that were indistinguishable in slices prepared from WT and KO mice (1.8 +/- 0.4 h and 2.1 +/- 0.2 h, respectively) and that were also sensitive to WAY-100,635. Unlike the pattern with 8-OH-DPAT, however, 5-HT-induced phase advances, in both WT and KO mice, were blocked by ritanserin, in this paradigm useful as a 5-HT(5A/7) antagonist (in addition to its more typical role as a 5-HT2A/2C antagonist). Serotonin antagonists when administered alone were without effect in slices from WT mice but produced significant phase shifts when administered to those from KO animals. Taken together, these results highlight the importance of the species used in establishing receptor mechanism. More provocatively, they support the involvement of multiple serotonin receptors in shifting the phase of circadian rhythms at ZT6.

Serotonin-induced phase advances of SCN neuronal firing in vitro: a possible role for 5-HT5A receptors?

Spontaneous firing rates of neurons in the suprachiasmatic nuclei (SCN) follow a consistent pattern, peaking near the midpoint of the light phase in a 12:12 light/dark schedule, and repeating this brief period of increased activity in subsequent circadian cycles. These carefully timed fluctuations reflect the output signal of the SCN, long recognized as the site of the endogenous biological clock in mammals. In rat hypothalamic slices, bath incubations of 8-OH-DPAT had previously been shown to elicit phase advances when applied at ZT6 (or 6 h following the onset of light), an action that could readily be attributed to 5-HT7 receptor activation. The present studies set out with the simple goal of establishing that the same receptor mechanism was responsible for the phase-shifting actions of 5-HT itself. Surprisingly, the phase advances elicited by 5-HT (0.5 microM, 1 h) at ZT6 were reduced by one 5-HT7 antagonist, ritanserin (10 microM), but not by another, mesulergine (10 microM). Receptor binding studies demonstrated a 25-fold greater affinity of ritanserin for h5-HT5A sites compared to mesulergine (Ki = 71 nM vs. 1,800 nM), an observation suggestive of a 5-HT5A mechanism for 5-HT and consistent with earlier observations of robust labeling of 5-HT5A sites in the SCN. 5-HT generated by the addition of L-tryptophan (10 microM, 1 h) to the slices displayed the same pattern of sensitivity, that is, blockade by ritanserin but not by mesulergine. Rp-cAMPS, a cAMP antagonist, failed to block the phase shifts elicited by 5-HT at a concentration (1 microM) previously shown to be effective against 8-OH-DPAT-induced phase shifts, in keeping with the proposed negative coupling of 5-HT5A receptors to cAMP production. Taken together, these results suggest that activation of both 5-HT5A and 5-HT7 receptors can produce phase advances of the circadian clock in vitro when they occur during mid-subjective day.

8-OH-DPAT as a 5-HT7 agonist: phase shifts of the circadian biological clock through increases in cAMP production.

Neurons in the suprachiasmatic nucleus (SCN), the site of the endogenous biological clock in mammals, fire spontaneously, peaking in firing rate near ZT6 or at the midpoint of the light phase in a 12:12 light-dark cycle. In rat hypothalamic slices, tissue incubations with drugs can produce a shift in this daily rhythm, the magnitude of which is dependent upon dose and the time of treatment. Previous work with 8-OH-DPAT had noted its ability to produce a phase advance, an earlier occurrence of the peak in neuronal firing, when applied at ZT6. Activation of 5-HT7 receptors was thought to be responsible for the shift, despite the clear preference of 8-OH-DPAT for 5-HT1A sites in terms of receptor binding affinity. In the present work, the actions of 8-OH-DPAT in SCN slices were confirmed and expanded to include additional dose-response and antagonist treatments. By itself, 8-OH-DPAT produced a concentration-dependent phase advance that was sensitive to co-application with 5-HT7 antagonists (ritanserin, mesulergine, SB-269970), but not to 5-HT1A antagonists (WAY-100,635, UH-301). Assignment of the receptor mechanisms for the antagonists employed was accomplished in experiments measuring binding affinities and the generation of cAMP, the latter monitored in a HEK-293 cell line expressing the r5-HT7 receptor and in tissue derived from rat SCN. The results indicate that the increases observed in cAMP levels are small but appear to be sufficient to produce a pharmacological resetting of the clock pacemaker. By aiding in the identification of the 5-HT receptor subtype responsible for the observed phase shifts and cAMP changes, 8-OH-DPAT represents an important pharmacological tool for 5-HT7 receptor activation, essentially broadening its role as the prototypical 5-HT1A agonist to one combining these two receptor activities.