Two mechanisms of quantized calcium release in skeletal muscle

Skeletal muscle uses voltage sensors in the transverse tubular membrane that are linked by protein-protein interactions to intracellular ryanodine receptors, which gate the release of calcium from the sarcoplasmic reticulum. Here we show, by using voltage-clamped single fibres and confocal imaging, that stochastic calcium-release events, visualized as Ca2+ sparks, occur in skeletal muscle and originate at the triad. Unitary triadic Ca(2+)-release events are initiated by the voltage sensor in a steeply voltage-dependent manner, or occur spontaneously by a mechanism independent of the voltage sensor. Large-amplitude events also occur during depolarization and consist of two or more unitary events. We propose a 'dual-control' model for discrete Ca2+ release events from the sacroplasmic reticulum that unifies diverse observations about Ca(2+)-signalling in frog skeletal muscle, and that may be applicable to other excitable cells.     

Comparison of autologous bone marrow transplantation with sequential chemotherapy for intermediate-grade and high-grade non-Hodgkin's lymphoma in first complete remission: a study of 464 patients. Groupe d'Etude des Lymphomes de l'Adulte

PURPOSE: Intensive chemotherapy followed by autotransplantation has given promising results in partially responding or sensitive relapsed patients with aggressive non-Hodgkin's lymphoma. In 1987, we designed a randomized study to evaluate the potential benefit of a high-dose regimen containing cyclophosphamide, carmustine, and etoposide (CBV) followed by autotransplantation over a consolidative sequential chemotherapy (ifosfamide, etoposide, asparaginase, and cytarabine) in patients in first complete remission with intermediate- and high-grade non-Hodgkin's lymphoma. PATIENTS AND METHODS: Patients were younger than 55 years and had at least one adverse prognostic factor. Induction treatment was that of the LNH84 protocol with an open randomization on the anthracycline. Patients in complete remission were further randomly assigned to receive either consolidation procedure. RESULTS: After induction treatment, 464 patients were assessable for the consolidation phase. With a median follow-up duration of 28 months, the 3-year disease-free survival rate was 52% (95% confidence interval, 45% to 59%) in the sequential chemotherapy arm and 59% (95% confidence interval, 52% to 66%) in the autologous transplant arm (P = .46, relative risk = 0.90). The 3-year survival rate did not differ between sequential chemotherapy and autotransplantation, at 71% (95% confidence interval, 64% to 78%) and 69% (95% confidence interval, 62% to 76%), respectively (P = .60, relative risk = 1.11). CONCLUSION: For such a subset of patients, consolidation with the CBV regimen followed by autologous bone marrow transplantation is not superior to sequential chemotherapy.     

Influence of tempering on the feeding value of rolled corn in finishing diets for feedlot cattle

Crossbred yearling steers (n = 125; 372 kg) were used in a 109-d finishing trial. Steers were fed an 88% concentrate diet containing 65% corn (DM basis) as 1) dry rolled corn (DRC); 2) tempered rolled corn (TRC), 43 mg surfactant (SarTemp)/kg corn; 3) TRC, 172 mg surfactant/kg corn; 4) TRC, 430 mg surfactant/kg corn; and 5) steam-flaked corn (SFC). Corn moisture was greater (3.5%, P < .01) for TRC than for DRC but less (10%, P < .05) than for SFC. Starch enzymatic reactivity was less for TRC than for either DRC (18%, P < .05) or SFC (42%, P < .01). Tempering increased the integrity of rolled corn and reduced the amount of particles less than 2 mm in diameter by 54% (P < .01). Steam flaking corn increased (P < .01) proportion (78%) of the grain having a particle size distribution of greater than 8 mm, as compared with TRC (25%) and DRC (3%). Compared with DRC, tempering enhanced (P < .10) ADG (9%), feed efficiency (5%), and dietary NE (3%). Daily weight gain was similar (P > .10) for TRC and SFC. Feed efficiency (P < .10) and dietary NE (P < .01) were greater (6%) for SFC than for TRC. There were no differences (P > .10) between DRC and TRC in ruminal and total tract digestion of OM, N, and starch, and in ruminal microbial efficiency. Ruminal digestion of OM decreased (linear effect, P < .05) and ruminal microbial efficiency increased (linear effect, P < .05) with increasing surfactant concentration. Ruminal digestion of OM and starch, and flow of nonammonia N to the small intestine were greater (31, 56, and 14%, respectively, P < .01) for SFC than for TRC. Postruminal and total tract digestion of OM, N, and starch, and dietary DE were greater (P < .01) for SFC than for TRC. We concluded that tempering corn will enhance animal performance. Increasing the concentration of surfactant used in tempering may enhance ruminal microbial efficiency and lean tissue growth.     

Bidirectional long-term modification of synaptic effectiveness in the adult and immature hippocampus

Previously we showed that delivering 900 pulses to the Schaffer collateral-CA1 pathway at 1-3 Hz causes a lasting depression of synaptic effectiveness that is input specific and dependent on NMDA receptor activation (Dudek and Bear, 1992a). Here we describe experiments aimed at further characterizing this homosynaptic long-term depression (LTD) and comparing it with long-term potentiation (LTP). To address the question of whether depressed synapses can still be potentiated and vice versa, LTP was saturated with repeated high-frequency tetani, and then LTD was induced with low-frequency stimulation (LFS). A second strong tetanus then restored the potentiation, indicating that the same synapses whose transmission had been depressed by LFS were capable of subsequently supporting potentiation. In a complementary experiment, LTD was induced first and then a strong high-frequency tetanus was delivered. We found that the resulting LTP achieved the same absolute magnitude as that observed in control slices that had received the high-frequency stimulation alone. Next, the postnatal development of LTD was investigated in slices prepared from rats at 6-35 d of age. The consequences of LFS were far more pronounced in slices from young rats. LTD following 900 pulses at 1 Hz measured -45 +/- 4% in CA1 of rats less than 2 weeks old as compared with -20 +/- 4 in animals at 5 weeks postnatal. It was also found that LTD precedes the developmental onset of LTP in CA1. Finally, we addressed the question of whether LTD could be saturated by repeated episodes of LFS in slices prepared from 3-week-old rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

High prevalence of mutations in the microtubule-associated protein tau in a population study of frontotemporal dementia in the Netherlands

We report here the large-scale purification of vacuolar (V0V1)-type Na+-ATPase from Enterococcus hirae achieved using column anion-exchange and gel filtration chromatographies; 32 mg of purified enzyme comprising nine subunits, A, B, C, D, E, F, G, I, and K, was obtained from 20 liter culture. This amount is 500-fold larger than that reported in the previous paper [Murata, T., Takase, K., Yamato, I., Igarashi, K., and Kakinuma, Y. (1997) J. Biol. Chem. 272, 24885-24890]. The purified enzyme shows a high specific activity of ATP hydrolysis (35.7 micromol Pi released/min/mg protein). ATP-driven 22Na+ uptake by reconstituted V0V1-proteoliposomes exhibited an apparent Kt value for Na+ of 40 microM, which is near the Km value (20 microM) for Na+ of the ATP hydrolytic activity. Denatured gel electrophoresis revealed that six subunits, A, B, C, D, E, and F, are releasable as the V1 subunit from the V0V1 complex by incubation with ethylenediaminetetraacetic acid; subunit G was not identified. The remaining V0-liposomes containing I and K subunits catalyzed Na+ uptake in response to potassium diffusion potential (Deltapsi, inside negative); the Kt value for Na+ of this reaction was estimated to be about 2 mM. Inhibition by N,N'-dicyclohexylcarbodiimide (DCCD) of the Na+-ATPase activity and Deltapsi-driven Na+ uptake by the V0-liposomes was prevented by the presence of Na+, suggesting that the Na+ binding site overlaps with the DCCD-reactive site.     

Localization of human BRCA1 and its loss in high-grade, non-inherited breast carcinomas

Although the link between the BRCA1 tumour-suppressor gene and hereditary breast and ovarian cancer is established, the role, if any, of BRCA1 in non-familial cancers is unclear. BRCA1 mutations are rare in sporadic cancers, but loss of BRCA1 resulting from reduced expression or incorrect subcellular localization is postulated to be important in non-familial breast and ovarian cancers. Epigenetic loss, however, has not received general acceptance due to controversy regarding the subcellular localization of BRCA1 proteins, reports of which have ranged from exclusively nuclear, to conditionally nuclear, to the ER/golgi, to cytoplasmic invaginations into the nucleus. In an attempt to resolve this issue, we have comprehensively characterized 19 anti-BRCA1 antibodies. These reagents detect a 220-kD protein localized in discrete nuclear foci in all epithelial cell lines, including those derived from breast malignancies. Immunohistochemical staining of human breast specimens also revealed BRCA1 nuclear foci in benign breast, invasive lobular cancers and low-grade ductal carcinomas. Conversely, BRCA1 expression was reduced or undetectable in the majority of high-grade, ductal carcinomas, suggesting that absence of BRCA1 may contribute to the pathogenesis of a significant percentage of sporadic breast cancers.     

The effects of ethanol on a measure of conditioned fear in mice

To develop an efficient topical delivery system for piroxicam using poloxamer gel formulation, physicochemical behavior of piroxicam in poloxamer was studied. The gelling property of poloxamer and the solubility of piroxicam in the poloxamer were investigated. The interaction between piroxicam and poloxamer was studied by x-ray diffractometry (XRD), infrared (IR) spectroscopy and differential thermal analysis (DTA) with a solid dispersion, coprecipitate, or physical mixture. Poloxamer 407 solutions showed the property of a gel when the concentration was higher than 15% (w/w) and poloxamer 407 increased the aqueous solubility of piroxicam by about 11-fold at the concentration of 22.5% (w/w). The results of XRD did not show the crystalline from of piroxicam in the solid dispersion and results of IR spectroscopic analysis showed an association between functional groups of piroxicam and poloxamer.     

Testicular suture: a significant risk factor for infertility among formerly cryptorchid men

BACKGROUND/PURPOSE: Although fertility is decreased after cryptorchidism, the importance of risk factors, including parenchymal testicular suture, is unknown. The aim of this study was to examine the relationship between parenchymal testicular suture and failure to conceive a child for 1 year or longer among formerly cryptorchid men. METHODS: Men who underwent orchidopexy between 1955 and 1972 at the Children's Hospital of Pittsburgh (n = 619) were surveyed by questionnaire and their medical records reviewed. Only the men who attempted to conceive a child (n = 387) are included. RESULTS: Logistic regression analysis determined significant risk factors for infertility. Testicular suture was strongly related to infertility (RR, 7.56; 95% CI, 1.66, 34.39) as were bilateral cryptorchidism (RR, 5.51; 95% CI, 1.58, 19.24), varicocele (RR, 4.72; 95% CI, 1.42, 15.75), hormone treatment before surgery (RR, 3.69; 95% CI, 1.22, 11.11), and partner conception problem (RR, 3.32; 95% CI, 1.11, 9.90). CONCLUSIONS: Testicular suture was a potent independent determinant of infertility among formerly cryptorchid men who have orchidopexy. Bilateral cryptorchidism, hormone treatment, varicocele, and partner conception problems also were associated with increased infertility.     

Bronchial obstruction and exhaled nitric oxide response during exercise in cold air

This study examines whether exhausting exercise in cold air induces bronchial obstruction and changes in exhaled [NO] and in exhaled NO output (V'NO). Thus, eight well-trained males performed two incremental exercise tests until exhaustion, followed by 5 min of recovery in temperate (22 degrees C) and cold (-10 degrees C) environments, at random. At -10 degrees C, they were dressed in warm clothes. Ventilation (V'E), oxygen consumption (V'O2), carbon dioxide production, cardiac frequency (fC), and [NO] and V'NO were measured continuously. Before and after each test, the subjects' maximal expiratory flow-volume curves and peak expiratory flow, forced expiratory volume in one second (FEV1) and forced expiratory flow at 25 (FEF25), 50 (FEF50) and 75% (FEF75) of forced vital capacity were determined. At -10 degrees C, significant decreases in FEV1 and FEF75 were observed after exercise. At rest and at the same submaximal intensity, V'O2, V'E and fC did not differ significantly. At rest and up to approximately 50% peak V'O2, [NO] and V'NO values were lower at -10 degrees C than at 22 degrees C. Thereafter, and during recovery, the V'NO response became similar at both -10 and 22 degrees C. This study confirms that considerable hyperpnoea in cold air causes a detectable airway obstruction. This airway cooling also induces an initial decrease in the exhaled NO response. Since endogenous NO-production is involved in bronchial dilation, it cannot be excluded that this lack of production may favour the appearance of airway obstruction.     

Interstitial cystitis

Nine genes belonging to the mouse Ly49 multigene family of natural killer cell receptors have been identified to date. Two of these genes, Ly49h and i, are very closely related to the well characterized Ly49c gene in the carbohydrate recognition domain. Here we show by Southern blotting that at least two additional new sequences exist in C57BL/6 mice that are also closely related to Ly49c in the carbohydrate recognition domain. Furthermore, in contrast to Ly49a, extensive variation in the arrangement and number of Ly49c-related genes in different mouse strains was observed. To characterize and localize the new Ly49c-related genes in C57BL/6 mice, we isolated and mapped genomic P1 clones hybridizing to an Ly49C exon 7 probe. Locations and the relative order of all Ly49 genes found within the clones was determined. We also used polymerase chain reaction to sequence exons 2, 4, and 7 from all genes. In this manner, we identified five new potential Ly49 genes which have been tentatively termed Ly49j-n. Ly49j, k, and n belong to the Ly49c-related subfamily, whereas Ly49l and Ly49m are most similar to Ly49d and g, respectively. Interestingly, the members of the Ly49c-related subfamily are not clustered as a unit but are interspersed among other Ly49 genes. These results illustrate the complex nature of the Ly49 gene family and should aid in the understanding of functions, such as the mediation of hybrid resistance, in which Ly49c-related genes play a role.