Kinetochore microtubules in crane-fly spermatocytes: untreated, 2 degrees C-treated, and 6 degrees C-grown spindles

We investigated the involvement of kinetochore microtubules (kMTs) in mediating chromosome-to-pole connections in crane-fly (Nephrotoma suturalis and Nephrotoma ferruginea) spermatocytes. Two experimental treatments were used to yield spindles with reduced numbers of nonkinetochore microtubules (nkMTs). Short-term (10-15 min) exposure of spermatocytes to 2 degrees C caused depolymerization of the majority of nkMTs, resulting in a kMT:(kMT + nkMT) ratio of 0.76. Long-term (24h) exposure to 2 degrees C followed by recovery at 6 degrees C resulted in a kMT:(kMT + nkMT) ratio of 0.55, the spindle having more nkMTs than a 2 degrees C-treated spindle but fewer than an untreated spindle, in which the kMT:(kMT + nkMT) ratio was 0.27. The numbers and lengths of kMTs in 6 degrees C-grown spindles were similar to those in untreated cells, suggesting that the overall inhibition of MT assembly at 6 degrees C apparently did not affect the mechanism by which kMTs are formed. We observed most kMTs of early anaphase spindles to be long (greater than 3 microns), and many extended to the polar regions of the spindle. Thus, the crane-fly spindle appears not to be as atypical as it was previously suggested to be.     

Post-mortem changes in cytochemical localization and enzymological measurement of marker enzymes of the mitochondria, SDH and Mg-ATPase, of porcine muscle stored at 4 degrees C, -18 degrees C, or -80 degrees C

As a series of studies on postmortem changes in the fine structure of porcine muscle, activity of two mitochondrial marker enzymes, succinate dehydrogenase (SDH) and magnesium dependent adenosine triphosphatase (Mg-ATPase), was measured and localized in cardiac, red and white muscles stored at 4 degrees C, -18 degrees C or -80 degrees C. The postmortem loss of SDH activity was most remarkable in cardiac muscle. The variation of SDH activity was proportional to the amount of absolute activity. The postmortem change of Mg-ATPase was more variable than SFH, though the activity was well preserved up to 15 weeks in all three types of porcine muscle stored at -80 degrees C. The loss of Mg-ATPase was most remarkable in red muscle stored at -18 degrees C or -80 degrees C. Cytochemical localization of SDH was between the outer and the inner mitochondrial membranes while that of Mg-ATPase was on the inner surface or matrix side of the inner membrane. Those localization was not altered by the difference in temperature and the duration of storage.     

Thermotolerance induced by heat, sodium arsenite, or puromycin: its inhibition and differences between 43 degrees C and 45 degrees C

When CHO cells were treated either for 10 min at 45-45.5 degrees C or for 1 hr with 100 microM sodium arsenite (ARS) or for 2 hr with 20 micrograms/ml puromycin (PUR-20), they became thermotolerant to a heat treatment at 45-45.5 degrees C administered 4-14 hr later, with thermotolerance ratios at 10(-3) isosurvival of 4-6, 2-3.2, and 1.7, respectively. These treatments caused an increase in synthesis of HSP families (70, 87, and 110 kDa) relative to total protein synthesis. However, for a given amount of thermotolerance, the ARS and PUR-20 treatments induced 4 times more synthesis than the heat treatment. This decreased effectiveness of the ARS treatment may occur because ARS has been reported to stimulate minimal redistribution of HSP-70 to the nucleus and nucleolus. Inhibiting protein synthesis with cycloheximide (CHM, 10 micrograms/ml) or PUR (100 micrograms/ml) after the initial treatments greatly inhibited thermotolerance to 45-45.5 degrees C in all cases. However, for a challenge at 43 degrees C, thermotolerance was inhibited only for the ARS and PUR-20 treatments. CHM did not suppress heat-induced thermotolerance to 43 degrees C, which was the same as heat protection observed when CHM was added before and during heating at 43 degrees C without the initial heat treatment. These differences between the initial treatments and between 43 and 45 degrees C may possibly be explained by reports that show that heat causes more redistribution of HSP-70 to the nucleus and nucleolus than ARS and that redistribution of HSP-70 can occur during heating at 42 degrees C with or without the presence of CHM. Heating cells at 43 degrees C for 5 hr after thermotolerance had developed induced additional thermotolerance, as measured with a challenge at 45 degrees C immediately after heating at 43 degrees C. Compared to the nonthermotolerant cells, thermotolerance ratios were 10 for the ARS treatment and 8.5 for the initial heat treatment. Adding CHM (10 micrograms/ml) or PUR (100 micrograms/ml) to inhibit protein synthesis during heating at 43 degrees C did not greatly reduce this additional thermotolerance. If, however, protein synthesis was inhibited between the initial heat treatment and heating at 43 degrees C, protein synthesis was required during 43 degrees C for the development of additional thermotolerance to 45 degrees C.(ABSTRACT TRUNCATED AT 400 WORDS)     

In vitro survival of Echinococcus granulosus protoscolices in several media, at +4 degrees C and +37 degrees C

As a first step towards hydatid disease control, the in vitro survival of protoscolices of Echinococcus granulosus was investigated for various media and temperatures. Higher percentage survival was obtained than previously reported: at 4 degrees C, 100% survival was obtained for 20 days in medium 199 (GIBCO) and for 25 days in hydatid fluid from the host of origin. Maximal survival was 30% at 55 days in these conditions. Flame cell activity was the criterion of choice for viability. At 37 degrees C survival rates were lower and morphological changes in protoscolices were observed.     

Thermal, metabolic, and cardiovascular responses to various degrees of cold stress.

The metabolic, thermal, and cardiovascular responses of two male Caucasians to 1 2 h exposure to ambient temperature ranging between 28 degrees C and 5 degrees C were studied and related to the respective ambient temperatures. The metabolic heat production increased linearly with decreasing ambient temperature, where heat production (kcal times m- minus 2 times h- minus 1) = minus 2.79 Ta degrees C + 103.4, r = -0.97, P smaller than 0.001. During all exposures below 28 degrees C, the rate of decrease in mean skin temperature (Tsk) was found to be an exponential function dependent upon the ambient temperature (Ta) and the time of exposure. Reestablishment of Tsk steady state occurred at 90-120 min of exposure, and the time needed to attain steady state was linearly related to decreasing Ta. The net result was that a constant ratio of 1.5 of the external thermal gradient to the internal thermal gradient was obtained, and at all experimental temperatures, the whole body heat transfer coefficient remained constant. Cardiac output was inversely related to decreasing Ta, where cardiac output (Q) = minus 0.25 Ta degrees C + 14.0, r = minus 0.92, P smaller than 0.01. However, the primary reason for the increased Q, the stroke output, was also described as a third-order polynomial, although the increasing stroke volume throughout the Ta range (28-5 degrees C) was linearly related to decreasing ambients. The non-linear response of this parameter which occurred at 20 degrees C larger than or equal to Ta larger than or equal to 10 degrees C suggested that the organism's cardiac output response was an integration of the depressed heart rate response and the increasing stroke output at these temperatures.     

Muscle glycogen depletion during exercise at 9 degrees C and 21 degrees C

This study compared glycogen depletion in active skeletal muscle after light and moderate exercise in both cold and comfortable ambient conditions. Twelve male subjects (Ss) were divided into two groups equally matched for the submaximal exercise intensity corresponding to a blood lactate concentration of 4 mM (W4) during cycle exercise. On two separate days Ss rested for 30 min at ambient temperatures of either 9 degrees C or 21 degrees C, with the order of temperature exposure being counter-balanced among Ss. Following rest a tissue specimen was obtained from the m. vastus lateralis with the needle biopsy technique. Six Ss then exercised on a cycle ergometer for 30 min at 30% W4 (range = 50 - 65 W) while the remaining group exercised at 60% W4 (range = 85 - 120 W). Another biopsy was taken immediately after exercise and both samples were assayed for glycogen content. Identical procedures were repeated for the second environmental exposure. No significant glycogen depletion was observed in the Ss exercising at 30% W4 in 21 degrees C, but a 23% decrease (p = 0.04) was observed when the same exercise was performed at 9 degrees C. A 22% decrease (p = 0.002) in glycogen occurred in the 60% W4 group at 21 degrees C, which was not significantly different from that observed during the same exercise at 9 degrees C. The results suggest that muscle substrate utilization is increased during light exercise in a cold environment as compared to similar exercise at a comfortable temperature, probably due to shivering thermogenesis.(ABSTRACT TRUNCATED AT 250 WORDS)