Cross-bridge kinetics studied with staircase shortening in single fibres from frog skeletal muscle

The kinetics of actin-myosin interaction has been studied in single active muscle fibres by repetitively eliciting tension transients with staircase shortening, consisting in a sequence of step releases of identical size (1--5 nm per half-sarcomere) imposed at regular time intervals (3--11 ms). Under sarcomere length-clamp conditions, the quick phase of tension recovery following each step in the staircase is the manifestation of the working stroke by synchronized cross-bridges. Different average shortening velocities are obtained by varying both the size of the step and the time interval between steps. Ti, the tension just before each step in the sequence, and T2, the tension attained at the end of the quick phase of tension recovery, decrease with the number of steps, reaching a steady state value, which is lower the larger the shortening velocity. In agreement with previous results on tension response to steady shortening, the overall shortening necessary to approach the steady state values of Ti and T2 is about 15 nm. The normalized amplitude of quick tension recovery (T2r), which is measured by the ratio of the amount of tension recovered at the end of the quick phase (T2--T1) over the tension drop simultaneous with the step (Ti--T1), has been used to measure the extent of the working stroke elicited by each step in the staircase. The steady state value of T2r decreases progressively with the increase of shortening velocity. At velocities higher than 0.5 μm s−1 per half-sarcomere the steady state value of T2r is attained after a transitory depression, which reaches a maximum for an amount of overall shortening increasing from about 8 nm up to about 13 nm with increase in shortening velocity from 0.5 to 1.4 μm s−1 per half-sarcomere. The velocity-dependent transitory depression of T2r can be explained with the mechanical-kinetic model described previously. In the model cross-bridges cycle through two pathways distinct for the kinetics of the detachment/reattachment process. Shortening promotes are distribution of cross-bridges interacting in the isometric conditions among the various states of the force-generating process. Shortening at high speed, preventing most of cross-bridges from undergoing the relatively fast (100 s−1) detachment/reattachment process, uncovers a rate limiting step in the cycle at the end of the 12 nm working stroke. Under these conditions, the finding that the fraction of the working stroke elicited by each step is transitorily depressed with respect to the steady state value reveals that in the original isometric state a large fraction of interacting cross-bridges was accumulated near the beginning of the working stroke This revised version was published online in July 2006 with corrections to the Cover Date.     

EFFECT OF SUBACUTE CABERGOLINE TREATMENT ON PROLACTIN, THYROID STIMULATING HORMONE AND GROWTH HORMONE RESPONSE TO SIMULTANEOUS ADMINISTRATION OF THYROTROPHIN-RELEASING HORMONE AND GROWTH HORMONE-RELEASING HORMONE IN HYPERPROLACTINAEMIC WOMEN

It is known that dopaminergic neurotransmission is involved in the control of PRL, TSH and GH secretion. Cabergoline (CAB) is a new ergolinic derivative with a long-acting dopaminergic activity. We evaluated 11 women with pathological hyperprolactinaemia before and during sub-acute CAB treatment (0.8-1.2 mg/p.o.; 8 weeks). Simultaneous administration of TRH (200 micrograms i.v.) and GHRH 1-44 (50 micrograms i.v.) were carried out before and after 4, 8 and 10 week intervals from the beginning of CAB treatment. Basal PRL levels (2453.5 +/- S.E. 444.5 mU/l) were significantly reduced during CAB administration (week 4: 164.5 +/- 66.5 mU/l; week 8: 168.0 +/- 66.5 mU/l; P less than 0.01) and no variations were observed 2 weeks after drug discontinuation (week 10: 210.0 +/- 98.0 mU/l). PRL percentage change after TRH was increased by CAB (P less than 0.05). No variation in basal and TRH-stimulated TSH levels was found during CAB administration. A slight increase in GH basal levels (3.0 +/- 0.6 mU/l) was found after weeks 4 (6.4 +/- 2.0 mU/l) and 10 (5.8 +/- 1.6 mU/l) (P less than 0.05). GH response to GHRH was significantly enhanced (ANOVA: P less than 0.01) during sub-acute CAB treatment. A positive correlation was found between GH secretory area and weeks of CAB therapy (P less than 0.01). Our data show that CAB is very effective in lowering PRL secretion in hyperprolactinaemia, and is able to modify PRL and GH responses after TRH and GHRH. The increasing trend in GH basal and GHRH-stimulated GH levels seems to indicate that CAB can override the central dopaminergic tone which is operative in hyperprolactinaemia.     

Insulin deficiency and insulin resistance in Type 2 (non-insulin-dependent) diabetes: quantitative contributions of pancreatic and peripheral responses to glucose homeostasis

A non-steady state dose-response study was designed to quantitate peripheral sensitivity to insulin and pancreatic responsiveness to glucose, and to assess their relative contribution to glucose intolerance in Type 2 diabetes (Type 2 DM, non-insulin-dependent). Eleven lean and eleven obese patients with mild diabetes (fasting plasma glucose, FPG, 10.3 +/- 1.0 and 9.4 +/- 0.6 mmol l-1, respectively) were examined; twenty-six lean and twelve weight-matched obese subjects served as controls. Pancreatic response was measured by sequential injection of 0.1, 0.3 and 0.9 g kg-1 glucose; peripheral sensitivity to insulin was determined from the rate of clearance (Kgluc) of 0.3 g glucose injected sequentially together with 25, 50 and 100 mU insulin kg-1 or with 0, 12.5 and 50 mU kg-1, under somatostatin infusion. The mean dose-response curve describing glucose-induced insulin release showed increased maximal capacity to secrete insulin in obese controls, while the responses of lean as well as obese Type 2 DM were reduced by more than 80%. The mean dose-response curves relating plasma exogenous insulin levels to Kgluc were similar in lean diabetics and lean controls. The curves of both obese controls and obese diabetics were shifted to the right, demonstrating similar insulin resistance. In four lean controls, sensitivity to insulin was tested also during a hyperglycemic clamp set at 10.3 +/- 0.6 mmol l-1. Hyperglycemia reduced the Kgluc at all insulin levels. Individual dose-response curves were transformed to single weighted numerical pancreatic responsiveness scores [PRS], and peripheral sensitivity scores [PSS].(ABSTRACT TRUNCATED AT 250 WORDS)     

Hybrid Therapies for Ventricular Arrhythmias

In recent years several trials demonstrated the efficacy of implantable cardioverter-defibrillation (ICD) therapy in reducing cardiac and total mortality in patients affected by rapid ventricular tachycardia (VT) and/or ventricular fibrillation. Nevertheless, ICD do not prevent arrhythmia recurrences, thus being a palliative and not a curative treatment modality. The tolerance to ICD therapy varies greatly, and within individuals, this leading to a nonuniform acceptance of this form of therapy. The very frequent occurrence of VT, defined as an arrhythmic storm, may be a life threatening condition. The majority of ICD patients is under antiarrhythmic drug therapy, to reduce episodes of VT or to make antitachycardia pacing more effective by slowing the tachycardia rate. Drug therapy, however, may cause additional problems, and does not represent the optimal solution. The prevention of VT and/or ventricular fibrillation episodes and excessive ICD therapy, remains a worthwhile goal. Radiofrequency catheter ablation (RFCA) is a curative approach, and can be expected to reduce the frequency of recurrent VT episodes in the majority of patients. The combination of these treatment modalities (ICD and RFCA) is often described as hybrid therapy, implying that the two treatments act providing some form of synergism. In experienced centers, RFCA is now performed, regardless of whether the VT rate is rapid and/or is hemodynamically unstable. Newer mapping and ablation techniques are now available, enhancing the acute success rate of the procedure. In this review the most recent application of VT catheter ablation and the use of advanced mapping and ablation techniques will be discussed.     

1 H-n.m.r. studies on the existence of substrate binding sites in bovine pancreatic ribonuclease A

The reaction of ribonuclease A with either 6-chloropurine riboside 5′-monophosphate or the corresponding nucleoside yields one derivative, with the reagent covalently bound to the α-amino group of Lys-1, called derivative II and derivative E, respectively. We studied by means of ¹H-n.m.r. at 270 MHz the interaction of these derivatives with different purine ligands. The pK values of His-12- and -119 were obtained and compared with those resulting from the interaction with ribonuclease A. The results showed that the interaction of derivative E with 3′AMP is similar to that described for RNase A as the pK₂ of His-12 is increased while that of His-119 remains unaltered. However, derivative II presents some differences as it was found an enhancement of the pK₂ values of both His-12 and His-119. Interaction of derivative II and derivative E with dApdA increases the pK₂ of His-119, whereas a decrease is found when it interacts with ribonuclease A. These results suggest that the phosphate group and the nucleoside of both derivatives are located in regions of the enzyme where natural substrate analogues have secondary interactions and they can be interpreted as additional binding sites.     

S-deoxo-Abu1, Ile3-amaninamide,an inactive amatoxin analogue

The title compound 3, an amatoxin analogue containing l -α-aminobutyric acid instead of l -asparagine in position 1, as in natural toad stool peptides, has been synthesized. It does not inhibit the eukaryotic DNA-dependent RNA polymerase form II (or B) in concentrations up to 10^-4M, whereas 50% inhibition is exerted in M solution by the corresponding Asn-analogue S-deoxo-Ile³-amaninamide 2. The striking difference seems to be due to a relatively small variation of the conformation recognized by sensitive NMR spectroscopic methods.     

Gender-Specific Effects of Prenatal Chlordane Exposure on Myeloid Cell Development

Work previously reported by this laboratory indicated that prenatalchlordane exposure affected macrophage function in young adultmice. Because these macrophage effects were due to exposureduring the development of the immune system, the possibilityof a persistent effect on the development of myeloid stem andprogenitor cells was considered. Female mice were treated witheither 0 or 8 mg of chlordane per kilogram body weight dailyfor 18 days during pregnancy. Myeloid hemopoietic activity ofbone marrow cells from 6-week-old offspring was evaluated forin vitro colony-forming units-in-culture in response to exogeneouslyadded recombinant forms of the cytokines granulocyte/macrophage-colonystimulating factor, macrophage- CSF, and interleukin 3 (IL-3).There was a significant depression of the numbers of bone marrowcolony forming units-granulocyte/macrophage (CFU-GM), CFU-IL-3,and CFU-macrophage (CFU-M) in only the female offspring. Maleoffspring consistently demonstrated no difference in the CFU-GM,CFU-IL-3, or CFU-M. Prenatal treatment with chlordane did notsignificantly affect the number of recoverable viable bone marrowcells in either male or female mice.     

Report of Three Cases with Tricho-Rhino-Phalangeal Syndrome Type I (Two Cases) and Type II (One Case)

Phenotypical and radiologic features of two unrelated patients with the syndrome Tricho- Rhino-Phalangeal syndrome type I (TRP I) and one with the syndrome TRP I1 are reported. All three cases were sporadic and the parents were non-consanguineous. The interstitial deletion of the chromosome 8q, evaluated on pro-. metaphases chromosomes, was not observed in any of the patients.     

DOES TESTOSTERONE INFLUENCE THE POST-STIMULATORY LEVELS OF CALCITONIN IN NORMAL MEN?

We have analysed the sex difference of calcitonin (CT) levels after combined stimulation with calcium and pentagastrin (Ca-PG) in the normal population, and the relationship of the post-stimulatory CT levels with free testosterone (FT). We have also studied the correlation between CT values and the anthropometric parameters, body mass index (BMI) and body surface area (BS), as well as the relationship between CT levels and calcium. A positive and statistically significant correlation was found between post-stimulatory CT and the increment over the base-line of CT and basal FT, and with the anthropometric parameters. However, the increment of CT and the peak values of CT did not have any significant correlation with the Ca levels (basal or post-stimulation). We conclude that the enhanced CT response found in normal men compared to normal women is at least partially determined by the higher testosterone levels found among normal men.