Acute morphine affects the rat circadian clock via rhythms of phosphorylated ERK1/2 and GSK3β kinases and Per1 expression in the rat suprachiasmatic nucleus

Background and Purpose

Opioids affect the circadian clock and may change the timing of many physiological processes. This study was undertaken to investigate the daily changes in sensitivity of the circadian pacemaker to an analgesic dose of morphine, and to uncover a possible interplay between circadian and opioid signalling.

Experimental Approach

A time-dependent effect of morphine (1 mg·kg⁻¹, i.p.) applied either during the day or during the early night was followed, and the levels of phosphorylated ERK1/2, GSK3β, c-Fos and Per genes were assessed by immunohistochemistry and in situ hybridization. The effect of morphine pretreatment on light-induced pERK and c-Fos was examined, and day/night difference in activity of opioid receptors was evaluated by [³⁵S]-GTPγS binding assay.

Key Results

Morphine stimulated a rise in pERK1/2 and pGSK3β levels in the suprachiasmatic nucleus (SCN) when applied during the day but significantly reduced both kinases when applied during the night. Morphine at night transiently induced Period1 but not Period2 in the SCN and did not attenuate the light-induced level of pERK1/2 and c-Fos in the SCN. The activity of all three principal opioid receptors was high during the day but decreased significantly at night, except for the δ receptor. Finally, we demonstrated daily profiles of pERK1/2 and pGSK3β levels in the rat ventrolateral and dorsomedial SCN.

Conclusions and Implications

Our data suggest that the phase-shifting effect of opioids may be mediated via post-translational modification of clock proteins by means of activated ERK1/2 and GSK3β.Tables of Links

Beta-blockade and exercise capacity in patients with mitral stenosis in sinus rhythm

BACKGROUND AND AIM OF THE STUDY: The study aim was to determine whether beta-blocker treatment (atenolol) improves cardiopulmonary exercise performance and ventilatory response in patients with mitral stenosis in sinus rhythm. METHODS: A prospective study comparing the results of cardiopulmonary exercise tests (CPETs) was performed before and after atenolol therapy in 17 patients in NYHA classes I and II with mitral stenosis in sinus rhythm. Transthoracic echocardiography was performed pre-study, and left ventricular diameters, ejection fraction and mitral valve area monitored. CPETs (Naughton protocol) were performed by two different investigators before and after one-week atenolol therapy (50 mg/day). The second investigator was blinded to the result of the baseline test. O2 consumption, CO2 production, ventilatory parameters and respiratory exchange ratios were measured on line. RESULTS: Maximal O2 uptake (VO2max) did not differ significantly before and after beta-blockade (median 16.8 and 15.0 ml/kg/min, respectively. Median heart rate at rest (72 versus 55 beats/min; p = 0.0003) and during peak exercise (153 versus 105 beats/min; p = 0.0003), and anaerobic threshold (10 versus 8.9 ml/kg/min; p = 0.02) were lower with beta-blockade compared with the baseline state. Minute ventilation at maximum exercise (41 versus 40 l/min) and ventilatory equivalent for CO2 (34 versus 35) were unchanged with atenolol therapy, indicating no improvement in ventilatory performance. When patients were grouped into those in whom VO2max was improved with atenolol therapy (n = 7) and those in whom it was impaired (n = 10), there were no inter-group differences with respect to age, left ventricular function, severity of mitral stenosis, NYHA class and grade of beta-blockade reached. Four patients felt symptomatically worse during atenolol treatment (lower NYHA functional class). CONCLUSION: Beta-blockade does not improve exercise tolerance in patients with mitral stenosis in sinus rhythm. In addition, ventilatory performance does not change with treatment.     

Intracytoplasmic sperm injection with cryopreserved testicular spermatozoa

To assess if testicular sperm cryopreservation is a valid alternative to repetition of testicular sperm retrieval techniques, results of a cryopreservation technique in pills have been retrospectively analyzed. Enough motile spermatozoa for ICSI were obtained in 172 from 190 (90.5%) frozen-thawed testicular sperm samples. Overall, 249 couples underwent 390 ICSI cycles, 156 using fresh and 234 using cryopreserved testicular sperm. Mean two-pronuclear fertilization rates per cycle were not significantly different after ICSI with fresh (62.0%) or with cryopreserved (63.2%) spermatozoa. Mean embryo cleavage rate per cycle was higher in the fresh (90.6%) than in the cryopreserved (84.6%) group (P = 0.016). However, clinical pregnancy rates per cycle (28.2% with fresh vs 27.8% with cryopreserved), implantation rates (12.2% vs 13.1%) and ongoing pregnancy rates per cycle (22.4% vs 21.8%) were not significantly different. Cryopreservation of testicular spermatozoa is an effective technique that can be used both in obstructive and in non-obstructive azoospermia.     

Systemic inflammation after inspiratory loading in chronic obstructive pulmonary disease

Objective

Patients with chronic obstructive pulmonary disease (COPD) present systemic inflammation. Strenuous resistive breathing induces systemic inflammation in healthy subjects. We hypothesized that the increased respiratory load that characterizes COPD can contribute to systemic inflammation in these patients.

Patients and methods

To test this hypothesis, we compared leukocyte numbers and levels of circulating cytokines (tumor necrosis factor alpha [TNFα], interleukin-1β [IL-1β], IL-6, IL-8, and IL-10), before and 1 hour after maximal incremental inspiratory loading in 13 patients with stable COPD (forced expiratory volume in one second [FEV₁] 29 ± 2.5% ref) and in 8 healthy sedentary subjects (FEV₁ 98 ± 5% ref).

Results

We found that: (1) at baseline, patients with COPD showed higher leukocyte counts and IL-8 levels than controls (p < 0.01); and, (2) one hour after maximal inspiratory loading these values were unchanged, except for IL-10, which increased in controls (p < 0.05) but not in patients with COPD.

Conclusions

This study confirms the presence of systemic inflammation in COPD, shows that maximal inspiratory loading does not increase the levels of pro-inflammatory cytokines (IL-1β, IL-8) in COPD patients or controls, but suggests that the former may be unable to mount an appropriate systemic anti-inflammatory response to exercise.     

Simple pressure measurement is not reliable in detection of access stenosis in native AV fistulas

Dynamic arterial and venous pressures (PA, PV) are used as the simplest tools to assess vascular access quality (VAQ). An increased PV over three consecutive dialyses is believed to indicate a stenosis, a rule devised for synthetic grafts (AVG) but not adequately validated for AV fistulas (AVF). In this study dynamic PV and static intra-access pressure (calculated by means of the simplified formula PIA=(PA+PV)/2) changes were evaluated in 46 accesses in which balloon angioplasty had to eventually be performed. The whole group consisted of 30 forearm AVF, 5 upper arm AVF and 11 AVG. Pressures were compared in each patient at a time of satisfactory access flow (QVA) and immediately before the angioplasty and pressure difference over that period (deltaPV, deltaPIA) evaluated. Despite a significant drop in QVA over the follow-up interval in both AVF and AVG, the mean deltaPV and deltaPIA in AVF were only several mm Hg and the chosen threshold limit of 20 mmHg was exceeded in approximately 10% of patients only. The results in the AVG group were, however, very different: The mean deltaPV and deltaPIA were close to 20 mmHg and almost 60% of patients in the AVG group exceeded this limit. Evaluation of PIA did not improve stenosis detection in either group. It is concluded that PV and/or PIA monitoring may be useful to detect a stenosis in AVG but not in AVF.     

Haematopoietic stem cell mobilization in patients with multiple myeloma--comparison of 3 variations of the stimulating regime

In the literature various regimes are described for successful collection of haematopoietic stem cells in patients with multiple myeloma. Most frequently cyclophosphamide is used, 5 g/m2 combined with different doses of haematopoietic growth factors. In our group the yields and tolerance of three stimulating regimes are compared: 1. cyclophosphamide 5 g/m2 and filgrastim (G-CSF) 5 micrograms/kg 2. cyclophosphamide 5 mg/m2 and filgrastim 10 micrograms/kg 3. cyclophosphamide 5 g/m2 and figrastim 5 micrograms/kg along with erythropoitin 50 IU/kg. Cyclophsphamide is administered always on the first day and the haematopoietic growth factors then from the third day till the end of collection of haematopoitic cells. In patients with multiple myeloma where only four cycles of VAD chemotherapy preceded and where radiotherapy of the axial skeleton was not used, optimal collection of haematopoietic stem cells was achieved after administration of cyclohosphamide 5 g/m2 with subsequent administration of 5 micrograms/kg G-CSF. By increasing the dose of G-CSF to 10 micrograms/kg or adding 50 IU/kg erthropoietin did not lead to a significant improvement of the tolerance and yield of this procedure.