Mobilization of circulating leucocyte and lymphocyte subpopulations during and after short, anaerobic exercise

A group of 11 healthy athletes [age, 27.4 (SD 6.7) years; body mass, 75.3 (SD 9.2) kg; height, 182 (SD 8) cm; maximal oxygen uptake, 58.0 (SD 9.9) ml.kg-1.min-1] conducted maximal exercise of 60-s duration on a cycle ergometer [mean exercise intensity, 520 (SD 72) W; maximal lactate concentration, 12.26 (SD 1.35) mmol.l-1]. Adrenaline and noradrenaline, and leucocyte subpopulations were measured flow cytometrically at rest, after 5-min warming up at 50% of each individual's anaerobic threshold (followed by 5-min rest), immediately after (0 min), 15 min, 30 min, and 1, 2, 4 and 24 h after exercise. Granulocytes showed two increases, the first at 15 min and, after return to pre-exercise values, the second more than 2 h after exercise. Eosinophils also increased at 15 min but decreased below pre-exercise values 2 h after exercise. Total lymphocytes and monocytes had their maximal increases at 0 min. Out of all lymphocyte subpopulations CD3-CD16/CD56(+)- and CD8+CD45RO--cells increased most and had their maximal cell counts at 0 min. The CD3(+)-, CD4+CD45RO(+)-, CD8+CD45RO(+)-, and CD19(+)- increased at 0 min, but had their maximum at 15 min. During the hours after exercise CD3-CD16/CD56(+)-, CD3+CD16/CD56(+)-, CD8+CD45RO(+)- and CD8+CD45RO--cells were responsible for the lymphocytopenia. The CD3(+)- and CD3-CD16/CD56(+)-cells were lower 24 h after exercise than before exercise. Adrenaline and noradrenaline increased during exercise. In conclusion, short anaerobic exercise led to a sequential mobilization of leucocyte subpopulations. The rapid increase of natural killer cells and monocytes may have been due to increased blood flow and catecholamine concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pulsatile secretion of gonadotrophins, ovarian steroids and ovarian oxytocin during prostaglandin-induced regression of the corpus luteum in the cow

A luteolytic dose (500 micrograms) of cloprostenol was given on Day 12 of the oestrous cycle to 5 heifers. Blood samples were collected simultaneously from the caudal vena cava and jugular vein at 5-20-min intervals from -6 to 0 (control period), 0 to 12 and 24 to 36 h after PG injection. Pulses of LH were secreted concomitantly with pulses of FSH during all sampling periods. However, during the control period separate FSH pulses were detected resulting in a shorter (P less than 0.01) interpulse interval for FSH than LH (93 versus 248 min). LH and FSH pulse frequencies increased (P less than 0.01) beginning 1-3 h after PG to interpulse intervals of 59 and 63 min, respectively, and continued to be maintained 24-36 h after PG. Concomitantly there was a 2-3-fold increase (P less than 0.01) in basal concentrations and pulse amplitude for LH (but not FSH). FSH basal concentrations and pulse amplitudes decreased (P less than 0.05) in 3 heifers 24-36 h after PG. Pulsatile secretion of oestradiol was observed at frequencies similar to LH during the periods 4-12 h (3 heifers) and 24-36 h (2 heifers) after PG, respectively, resulting in higher (P less than 0.05) mean oestradiol concentrations. Progesterone concentrations in the vena cava increased (P less than 0.01) 5-10 min after PG but decreased (P less than 0.01) 67% by 20 min after PG. This decrease was followed by a rise (P less than 0.05) beginning 2-3 h after PG and lasting for an average of 3.3 h.(ABSTRACT TRUNCATED AT 400 WORDS)     

Synthesis,pharmacokinetics, and metabolism of the <Emphasis Type="Italic">C</Emphasis>-terminal tripeptide of dermorphin and its diastereomer

A tritium-labeled C-terminal fragment of dermorphin (H-Tyr-[3,4-³H]Pro-Ser-NH₂) and its isomer (H-Tyr-D-[3,4-³H]Pro-Ser-NH₂) with molar radioactivity of 35 Ci/mmol were synthesized, and their pharmacokinetics and metabolism in rat organs were studied after their intramuscular injections. The tripeptides were detected in the blood only for 5 min after the injection, and maximum contents of both compounds (approximately 5% of the total amount of the injected label) were registered in the kidneys after 20 min. Both stereomers were shown to penetrate into the brain. We failed to detect any radioactive metabolite, except proline, due to rapid proteolytic degradation of these peptides.     

Successful in vitro and in vivo development of in vitro fertilized two- to four-cell cat embryos following cryopreservation, culture and transfer

In vitro and in vivo survival of in vitro-derived 2- to 4-cell cat embryos following cryopreservation was examined. Prefreeze 1- vs 2-step cryoprotectant exposure (Experiment 1) and warming method (Experiment 2) on zona pellucida damage and development in vitro were compared. To determine viability in vivo, frozen/thawed embryos were cultured in vitro to the morula/early blastocyst stage and transferred to synchronous recipients (Experiment 3). At 24 to 26 h after IVF, embryos were cryopreserved in 1.4 M propanediol (Pr) + 0.125 M sucrose (Su) by cooling at 0.3 degrees C/min from -6 degrees C to -30 degrees C and storing in liquid nitrogen. Autologous embryos were cultured in vitro for 7 d. After warming for 5 sec in air and 10 sec at 37 degrees C in water (Experiments 1 to 3), or at room temperature air (22 degrees C; Experiment 2), the cryoprotectant was removed and embryos were cultured in vitro for 6 d (Experiments 1 and 2). Development was assessed after staining by counting cell numbers/embryo and determining the percentages at the 2- to 4-cell (nonsurvivor), pre (5 to 15), early (16 to 32), mid (33 to 50), late (>50) morula or blastocyst stages. Post-thaw development to late morula/blastocyst after 1-step exposure (68%, 15 min Pr + Su) was higher (P< 0.05) than that after 2-step exposure (36%, 15 min Pr and 15 min Pr + Su). Both warming methods produced similar percentages of embryos with damaged zonae (13 to 15%) and equivalent development to morula/blastocyst (64 to 69%). Development in vitro to early morula/blastocyst of frozen embryos with intact zonae was similar to that of nonfrozen embryos. Following cryopreservation, most 2- to 4-cell cat embryos retained their capability for in vitro development to morula/blastocyst, and in vivo viability was demonstrated by the birth of 3 live kittens to 2 of 4 recipients following the transfer of 58 embryos.     

Development of in vitro matured,in vitro fertilized domestic cat embryos following cryopreservation,culture and transfer

The ability of embryos to successfully survive cryopreservation is dependent on both morphological and developmental characteristics. Domestic cat oocytes matured in vitro exhibit alterations in nuclear and cytoplasmic maturation that may affect developmental competence, particularly after cryopreservation. In Experiment 1, we evaluated the developmental competence of in vitro produced (IVM/IVF) cat embryos after cryopreservation on Days 2, 4 or 5 of IVC. In Experiment 2, in vivo viability was examined by transfer of cryopreserved embryos into recipient queens. Oocytes recovered from minced ovaries were cultured in TCM-199 with hCG/eCG and EGF at 38 degrees C in 5% O(2), 5% CO(2), 90% N(2) for 24h. In Experiment 1, after IVM/IVF, on Day 2 (n=56), Day 4 (n=48) and Day 5 (n=42) of IVC, embryos were equilibrated for 10 min at 22 degrees C in HEPES (15m M) Tyrode's (HeTy) with 1.4M propylene glycol (PG), 0.125 M sucrose (S), 10% dextran and 10% FBS, loaded into 0.25 ml straws, cooled at 2.0 degrees C/min to -6.0 degrees C and held for 10 min. After seeding, cooling resumed at 0.3 degrees C/min to -30 degrees C and after a 10 min hold, straws were plunged into liquid nitrogen (LN(2)). Straws were thawed in air for 2 min and cryoprotectant was removed by a five-step rinse consisting of 3 min each in HeTY with 0.95 M PG/0.25 M S; 0.95 M PG/0.125 M S; 0.45 M PG/0.125 M S; 0 PG/0.125 M S; 0 PG/0.0625 M S. Contemporary IVM/IVF embryos were used as nonfrozen controls (Day 2, n=14; Day 4, n=26; Day 5, n=35). After 8 days of IVC, the number of embryos developing to blastocysts was recorded and blastocyst cell numbers were counted after staining with Hoechst 33342. In Experiment 1, developmental stage did not affect the survival rate after thawing (Day 2=79%, Day 4=90%, Day 5=98%) and was not different from that of controls (Day 2=89%, Day 4=88%, Day 5=96%). Blastocyst development was similar among days both after cryopreservation (Day 2=59%, Day 4=54%, Day 5=63%) and in controls (Day 2=55%, Day 4=54%, Day 5=58%). Mean (+/-S.D.) cell number of blastocysts was slightly lower (NS) in cryopreserved embryos (Day 2=152+/-19, Day 4=124+/-20, Day 5=121+/-24) than in controls (Day 2=141+/-25, Day 4=169+/-21, Day 5=172+/-19). In Experiment 2, embryos frozen on Day 2 (n=68), Day 4 (n=49) or Day 5 (n=73) were thawed and cultured for 3, 1, or 0 days before transfer by laparotomy to 5 (mean=12.6+/-2.4), 4 (mean=12.2+/-3.7) and 6 (mean=12.0+/-1.6) recipients, respectively. Four recipients were pregnant on Day 21; two from embryos frozen on Day 4 and two from Day 5. Two live kittens were born at 66 days, a third kitten died during parturition at 64 days and a fourth pregnancy aborted by Day 45. In summary, we have shown that a controlled rate cryopreservation technique can be successfully applied to cat embryos produced by IVM/IVF. In vitro development to the blastocyst stage was not affected by the age of embryos at cryopreservation. The births of live kittens after ET of cryopreserved embryos is additional validation of progress toward applying assisted reproductive technology to preservation of endangered felids.     

Output, Tissue Levels, and Synthesis of Acetylcholine During and After Transient Forebrain Ischemia in the Rat

Biochemical changes in the rat brain cholinergic system during and after 60 min of ischemia were studied using a four-vessel occlusion model. Extracellular acetylcholine (ACh) concentrations in the unanesthetized rat hippocampus markedly increased during ischemia and reached a peak (about 13.5 times baseline levels) at 5-10 min after the onset of ischemia. At 2-5 h after reperfusion, extracellular ACh concentrations were reduced to 64-72% of the levels of controls. ACh levels in the hippocampus, striatum, and cortex decreased significantly during ischemia and exceeded their control values just after reperfusion. A significant increase in hippocampal ACh level after 2 days of reperfusion and a decrease in [14C]ACh synthesis from [14C]glucose in hippocampal slices excised at 2 days after reperfusion were observed. The extracellular concentrations and tissue levels of choline markedly increased after ischemia. These results show that ACh is markedly released into the extracellular space in the hippocampus during ischemia, and they suggest that ACh synthesis is activated just after reperfusion and that cholinergic activity is reduced after 2-48 h of reperfusion in the hippocampus.     

Effect of suckling on cortisol, progesterone and luteinizing hormone in postpartum beef cows

Five primiparous, 3-year-old Hereford cows suckled ad libitum , were cannulated via the jugular vein and stanchioned for 2-day sampling periods, every 14 days starting 14 days after the mean calving date. On the second day of each period, calves were removed to a pen away from the cows, for 9 hours. Blood was sampled 5 min before calves were returned to their dams, as soon as possible after initiation of suckling (IOS), and at 15-min intervals for 45 min, thereafter. Cortisol, progesterone and luteinizing hormone (LH) concentrations in the serum were quantitated by radioimmunoassay. Mean serum cortisol concentrations were 7.3 +/- .7, 9.4 +/- .7, 12.1 +/- .9, 7.5 +/- .5 and 5.7 +/- .4 ng/ml (mean +/- S.E.) at -5, 0, 15, 30 and 45 min after IOS, respectively, for all cows across all periods. Cortisol concentrations, during and after suckling, tended (P<.06) to differ among sampling periods, during the postpartum interval. Serum progesterone concentrations were .28 +/- .02, .28 +/- .02, .32 +/- .05 and .24 +/- .03 ng/ml at 0, 15, 30 and 45 min after IOS, respectively, for all cows across all period, indicating that suckling had no effect on serum progesterone, and were similar at all sampling periods during the postpartum interval. Serum LH concentrations were .81 +/- .07, .77 +/- .06, .71 +/- .04, and .72 +/- .04 ng/ml at 0, 15, 30 and 45 min after IOS, respectively. During the postpartum interval, serum LH concentrations were greater (P<.01) at 71 and 85 days postpartum than at any other time.     

Interrelations of hypocapnia, hypoxia, cerebral blood flow and electrical activity of the brain under voluntary hyperventilation in humans

Changes of different physiological parameters in human caused by hyperventilation of 3-min and longer duration were investigated and correlated. It was found that during 3-min hyperventilation, resulting in 4.5-5 fold increase of the respiration velocity, similar phasing changes of the central and cerebral haemodynamics occurred. The blood flow velocity according to the rheographic data during the hyperventilation first increases, reaching maximum at 1st - 2nd min of the test, and then decreases, reaching minimum at 2nd - 3rd min after it's end, and then slowly increases. Cerebral blood flow velocity during all the 3 min of the hyperventilation in most of the subjects keeps being increased, and after the test - decreased. At the same time transcutaneous pressure of carbon dioxide changes differently - decreases to minimum (approximately 25 mmHg) at the end of the test and then increases, reaching approximately 90% of the background level, at 5th min after the end of the test. Oxygen saturation of the blood during the test is found to be 98-100% and decreases to 90% at 5th min after it's end, which in overall with cerebral blood flow decrease appears to be the factor of the brain's hypoxia. In different subjects "mirror" changes of the EEG spectral power of different EEG ranges in relation to transcutaneous pressure of carbon dioxide dynamics were revealed by the hyperventilation. Taking into account the factors of duration or recurrence of the hyperventilation is important for the understanding the interrelations of cerebral haemodynamics, hypocapnia, hypoxia and electrical activity of the brain. It was found that after the recurrent hyperventilation of increasing amount (several times in hour by 3 min) cerebral blood flow might decrease markedly against the background of relatively small changes of electrical activity of the brain. The discussing of the data presented in the paper is carried out from the point of view of important role of tissue oxygen utilization mechanisms of the brain in adaptation to hypoxia and hypocapnia.     

Effects of short-term stress, xylazine tranquilization and anesthetization with xylazine plus ketamine on plasma concentrations of cortisol, luteinizing hormone, follicle stimulating hormone and prolactin in ovariectomized pony mares

Long-term ovariectomized pony mares were subjected to one of four treatments: 1) control group - no treatment, 2) stressed group - 5 min of restraint via a twitch, 3) tranquilized group - administered xylazine (1.1 mg i.v. per kg of body weight), and 4) anesthetized group - administered xylazine followed 2 min later by ketamine (2.2 mg i.v. per kg of body weight). Blood samples were taken at -40, -30, -20, -10, -0.5, 10, 20, 30, 40, 50, 60 and 90 min and at 2, 3, 4, 6, 8 and 24 h relative to onset of treatment. Stress increased (P<0.05) cortisol concentrations 20 to 50 min after treatment and again at 6 and 8 h. Tranquilization had no effect on cortisol concentrations, whereas anesthetization increased (P<0.05) cortisol concentrations from 90 min through 8 h after treatment. Concentrations of luteinizing hormone (LH) and follicle stimulating hormone (FSH) did not vary (P>0.1) relative to pretreatment in any group of mares. Concentrations of prolactin were 2.7-fold higher (P<0.05) 24 h after treatment in all four groups, indicating some procedural or environmental influence on prolactin secretion. There was a transient increase (P<0.06) in prolactin concentrations in anesthetized mares 30 min after treatment. Although two of these three commonly used methods of restraint did affect cortisol concentrations, there was no effect on plasma concentrations of LH or FSH. Thus, we conclude that such methods of restraint can be used in short-term situations without disturbing estimates of LH and FSH secretion. However, when prolactin concentrations are to be measured, anesthesia with ketamine should not be used.     

Design, syntheses, and antitumor activity of novel chromone and aurone derivatives

A series of new chromone analogues bearing heterocyclic thioether moiety and aurone analogues bearing cyclic tertiary amine moiety were designed and synthesized under microwave irradiation. The synthetic protocol was found to present many advantages, such as higher yields, shorter reaction time (10-20 min), mild condition, and readily isolation of the products. The synthesized compounds were assayed for their antitumor activity against four kinds of human solid tumor cell lines including HCCLM-7, Hep-2, MDA-MB-435S, and SW-480. Two compounds, (Z)-2-((4-benzyl-piperazin-1-yl)methylene)benzofuran-3(2H)-one 5e and (Z)-2-((4-(bis(4-fluorophenyl)methyl)piperazin-1-yl)methylene)benzofuran-3(2H)-one 5f, were identified as the most promising candidates with the IC(50) values in the range of 4.1-13.1 microM. Further cell cycle studies revealed that compounds 5e and 5f arrest the cell cycle in G(0)/G(1) phase and displayed apoptosis-inducing effect on Hep-2 cells.     

Endocrine control of TAG lipase in the fat body of the migratory locust, Locusta migratoria

Aspects of the role and activation of the enzyme triacylglycerol lipase (TAG lipase) in the fat body of the migratory locust Locusta migratoria were investigated. TAG lipase is under the hormonal control of the three endogenous adipokinetic peptides of the migratory locust, Locmi-AKH-I, Locmi-AKH-II and Locmi-AKH-III. Injection of low doses (5-10 pmol) of each peptide causes an increase in lipase activity. The activation of lipase is time dependent: an elevated activity was recorded 15 min after injection of 10 pmol Locmi-AKH-I and maximum activation was reached after 45-60 min. The activation of TAG lipase is also dose-dependent. Doses of 2 pmol of each Locmi-AKH had no effect, whereas 5 pmol caused a significant activation. Maximum activation is reached with a dose of 10 pmol. Analogues of the second messengers cAMP (cpt-cAMP) and IP(3) (F-IP(3)) both activate the enzyme glycogen phosphorylase whereas only cpt-cAMP, but not F-IP(3), activates TAG lipase; cpt-cAMP elevates the lipid levels in the haemolymph. Activation of lipase is specific to the three endogenous AKH peptides: 5 pmol of the endogenous peptide Locmi-HrTH and 10 pmol of corazonin failed to activate lipase. High doses of octopamine did not activate lipase nor did they elevate the lipid concentration in the haemolymph. TAG lipase is stimulated by flight activity but activation is slower than that of glycogen phosphorylase: after 30 min of flight or after 5 min of flight plus 1h of subsequent rest, activity of TAG lipase is increased, but not immediately after 5 min of flight. In contrast, glycogen phosphorylase is activated significantly after 5 min of flight. These activation patterns of the two enzymes mirror-image the concentration of their substrates in the haemolymph: there is a significant decrease in the concentration of carbohydrates after 5 min of flight, whereas no change of the concentration of lipids can be measured after such short time of flight activity; however, a subsequent rest period of 1h is sufficient to increase the lipid concentration.     

Stimulation of FA and phosphatase-1 activities by insulin in 3T3-L1 cells

The phosphatase-1 activator FA and phosphatase-1 were assayed in 3T3-L1 cells exposed to insulin. The cytosolic FA activity was transiently stimulated (7-8-fold) 1 and 2 min after exposure to 10(-8) M insulin and returned to control values within 5-10 min. Cytosolic phosphatase-1 (assayed after trypsin treatment) was activated (120-140% of controls) between 2 and 5 min and returned to control values within 10 min. Insulin effects were dose-dependent, with maximum stimulation of both activities at 10(-8) M insulin. The possibility that FA and other kinases mediate phosphatase activation by insulin is discussed.     

Kinetics of phospholipase A2, arachidonic acid, and eicosanoid appearance in mouse zymosan peritonitis

Intraperitoneal injection of zymosan into mice induces a peritonitis characterized by cellular influx, plasma leakage and the appearance of arachidonic acid (AA) metabolites. We report that zymosan injection also stimulates the accumulation of AA, docosahexaenoic acid, linoleic acid, and phospholipase A2 (PLA2) activity. The amount of the unsaturated fatty acids (UnFA) varies both with the zymosan dose and time. Significantly increased levels of UnFA were first detected 15 min after zymosan injection. Maximal levels of the UnFA were reached 1 to 2 h post zymosan injection (AA: 725 +/- 29 ng/mouse, docosahexaenoic acid: 296 +/- 23 ng/mouse, linoleic acid: 4489 +/- 179 ng/mouse) and declined to saline control levels by 8 h. PLA2 activity was significantly increased 5 to 15 min after zymosan injection. Maximal levels of PLA2 activity occurred 15 to 30 min after zymosan injection (31.8 +/- 9.1 nmol phospholipid/mg protein/h) and then decreased by 30% through 24 h. Neither the appearance of UnFA nor PLA2 activity correlated with cellular influx, but both were coincident with plasma exudation at 5 to 15 min after zymosan. However, maximal exudation occurred 1 to 2 h post zymosan injection similar to that seen with the UnFA but not PLA2. These latter results suggest that a significant portion of the UnFA found in the peritoneal cavity of zymosan-injected mice originates from the plasma. PLA2 activity at the early time points (5 to 15 min) may also contribute to the levels of UnFA via hydrolysis of tissue and/or cellular phospholipids.     

Cryopreservation of channel catfish sperm: effects of cryoprotectant exposure time, cooling rate, thawing conditions, and male-to-male variation

The purpose of this study was to extend previous work on the cryopreservation of channel catfish (Ictalurus punctatus) sperm. The objectives were to compare the effects of freezing and thawing on motility of sperm for: (1) 1 or 48-h exposure before freezing to 5% methanol and use of 0.5 or 0.25 mL straws; (2) 1 h or 5-day exposure before freezing to 5% methanol; (3) cooling at 45 or 3 degrees C/min; (4) thawing at 30, 40 or 50 degrees C using 5 or 10 s duration, and (5) cryopreservation with 5 or 10% methanol of samples from 50 males to analyze male-to-male variation. No differences were found in motility reduction for 1 or 48 h exposure times in 5% methanol, for use of 0.5 or 0.25 mL straws, or for 1 h or 5-day exposures in 5% methanol. A cooling rate of 45 degrees C/min resulted in lower motility reduction (33+/-9%) than a rate of 3 degrees C/min (83+/-13%) (P=0.002). A thawing temperature of 50 degrees C resulted in lower motility reduction (25+/-14%) than 30 degrees C (51+/-21%) or 40 degrees C (59+/-11%) (P=0.001). A thawing duration of 10 s resulted in lower motility reduction (38+/-12%) than a duration of 5 s (52+/-12%) (P=0.005), and there was an interaction between thawing temperature and duration (P=0.050). A concentration of 5% methanol resulted in lower motility reduction (43+/-17%) than 10% methanol (67+/-14%) (P=0.001). Regression analysis showed no relationship between motility before freezing and after thawing for 5% methanol (r2=0.012) or 10% methanol (r2=0.011).     

Sperm cryopreservation of African catfish, Clarias gariepinus: cryoprotectants, freezing rates and sperm:egg dilution ratio

Methods for cryopreserving spermatozoa and optimizing sperm:egg dilution ratio in African catfish Clarias gariepinus were developed. Five percent to 25% DMSO and methanol were tested as cryoprotectants, by diluting semen in Ginzburg fish ringer and freezing in 1-milliliter cryovials in a programmable freezer. To avoid an excess of spermatozoa per egg, post-thaw semen was diluted 1:20, 1:200 or 1:2,000 before fertilization. Highest hatching rates were obtained by spermatozoa frozen in 10% methanol and post-thaw diluted to 1:200. Then, slow freezing rates (-2, -5 or -10 degrees C/min) to various endpoint temperatures (range -25 to -70 degrees C) before fast freezing in liquid nitrogen (LN2) were evaluated. Hatching rates equal to control (P > 0.05) were obtained by spermatozoa frozen at -5 degrees C/min to -45 to -50 degrees C and at -10 degrees C/min to -55 degrees C. In 3-step freezing programs, at -5 degrees C/min, the effect of holding spermatozoa for 0, 2 or 5 min at -30, -35 or -40 degrees C before fast freezing in LN2 was analyzed. Hatching rates equal to control (P > 0.05) were produced by spermatozoa frozen to, and held at, -35 degrees C for 5 min and at -40 degrees C for 2 or 5 min. Finally, frozen spermatozoa (10% methanol, -5 degrees C/min, 5-min hold at -40 degrees C, LN2, post-thaw diluted to 1:200) were tested in on-farm fertilization conditions. Again, no difference (P > 0.05) in hatching rate was observed between frozen and fresh spermatozoa. Cryopreservation offers utility as a routine method of sperm storage and management for catfish.     

Temperature, evapotranspiration and primary photochemical responses of apple leaves to hail

The objective of this work was to examine immediate physiological plant responses to hail and subsequent recovery in terms of evapotranspiration, leaf temperature and primary photochemical processes using apple as a model crop. Thermal emission pictures were taken in darkness to avoid interference from stomatal movements; temperature gradients were identified in concentric rings around sites of hail injury, with a distinct drop in temperature of up to 2.3 degrees C in the center immediately after the induced hail injury. This was due to enhanced evapotranspiration from the injured tissue. Six to twelve minutes after hail injury, the initial decrease in leaf temperature partially reversed. Chlorophyll fluorescence kinetics of light-adapted leaves showed a dramatic decrease in effective photosynthetic electron transport rate (ETR), from 20.5 to 9.0 micromol electron m(-2)s(-1) within 5 min from hail injury, and a rapid recovery to 14.1 micromol electron m(-2)s(-1) within the next 5 min. After 7h, ETR partially recovered to 17.4 micromol electron m(-2)s(-1). An initial drop in non-photochemical efficiency (NPQ) from 1.07 to 0.90 units within 5 min after hail injury was followed by a sharp increase to 1.67 units after another 5 min. During the next hour, NPQ gradually decreased to the initial level. This indicates increased thermal dissipation in photosystem II (PS II) as a protective mechanism against incident excessive energy in the leaves with closed stomata for 1h after hail injury. In contrast to the fluorescence kinetics of light-adapted leaves, maximum quantum yield Fv/Fm of PSII in the dark-adapted state remained unchanged at 0.79-0.81 relative units for the first 5 min after hail injury. Thereafter, Fv/Fm slowly declined to 0.75 within 1h, and to a trough of 0.73 at 3h. Seven hours after hail injury, Fv/Fm values were at 0.76, indicating partial recovery of PS II efficiency. The discrepancy in the dynamics of ETR and Fv/Fm responses may be explained by the formation of alternative electron sinks such as reactive oxygen species, particularly superoxides, which withdraw electrons from the photosynthetic transport, resulting in apparently higher values of calculated ETR.     

Evaluation of the pituitary-gonadal response to GnRH, and adrenal status, in the leopard (Panthera pardus japonensis) and tiger (Panthera tigris)

Frequent blood samples were collected to study hormonal responses to GnRH in male and female leopards and tigers. Animals were anaesthetized with ketamine-HCl and blood samples were collected every 5 min for 15 min before and 160 min after i.v. administration of GnRH (1 micrograms/kg body weight) or saline. No differences in serum cortisol concentrations were observed between sexes within species, but mean cortisol was 2-fold greater in leopards than tigers. GnRH induced a rapid rise in LH in all animals (18.3 +/- 0.9 min to peak). Net LH peak height above pretreatment levels was 3-fold greater in males than conspecific females and was also greater in tigers than leopards. Serum FSH increased after GnRH, although the magnitude of response was less than that observed for LH. Basal LH and FSH and GnRH-stimulated FSH concentrations were not influenced by sex or species. Serum testosterone increased within 30-40 min after GnRH in 3/3 leopard and 1/3 tiger males. Basal testosterone was 3-fold greater in tiger than leopard males. LH pulses (1-2 pulses/3 h) were detected in 60% of saline-treated animals, suggesting pulsatile gonadotrophin secretion; however, in males concomitant testosterone pulses were not observed. These results indicate that there are marked sex and species differences in basal and GnRH-stimulated hormonal responses between felids of the genus Panthera which may be related to differences in adrenal activity.     

The Influence of Bicuculline-Induced Seizures on Free Fatty Acid Concentrations in Cerebral Cortex, Hippocampus, and Cerebellum

Abstract: Using ventilated rats maintained on N₂O-O₂ (70:30, vol/vol) we induced continuous seizures with i.v. bicuculline and analysed free fatty acids (FFA) in cerebral cortex, hippocampus, and cerebellum after seizure durations of 1–120 min. In the cerebral cortex, peak FFA concentrations were observed after 5 min, with a threefold increase in total FFA content. The values then remained unchanged for the next 15-20 min, but decreased thereafter. At 60 and 120 min, total FFA contents were only moderately increased above control. In the initial period, arachidonic acid increased about 10-fold and stearic acid 2- to 3-fold, with little change in palmitic acid and linoleic acid concentrations. At all times, the docosahexenoic acid concentration was markedly increased. Following its massive accumulation at 1 min, arachidonic acid gradually decreased in concentration. Pretreatment of animals with indomethacin did not alter this behaviour. After 20 and 120 min of seizure activity, changes in total and individual FFA concentrations in the hippocampus were similar to those observed in the cerebral cortex. The cerebellum behaved differently. Thus, at 20 min the only significant change was a 5- to 10-fold increase in arachidonic acid concentration and, after 120 min, total and individual FFA concentrations were similar to control values. Furthermore, since the control values for arachidonic acid were much lower in the cerebellum, the 20-min values were only about 20% of those observed in the cerebral cortex and the hippocampus.     

Sensitivity of human pancreatic adenocarcinoma tumor lines to chemotherapy, radiotherapy, and hyperthermia

BACKGROUND: Surgical treatment of pancreatic adenocarcinoma has failed to produce many cures secondary to high rates of intraperitoneal relapses and liver metastases. The aim of this ex vivo study was to evaluate the inherent chemosensitivity, radiosensitivity and hyperthermic sensitivity of pancreatic adenocarcinoma and to investigate the usefulness of a 3-(4,5-dimetylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay utilized in each sensitivity test. METHODS: Nine human pancreatic adenocarcinomas were tested ex vivo after growth in nude mice. After 72 hr of chemotherapy, radiotherapy and hyperthermia, efficacy was assessed using MTT assay to determine the ratio of surviving fraction of treated cells-to-that of untreated control cells (TIC ratio). RESULTS: Tumor sensitivities as measured by the IC50 (drug concentration producing 50% growth inhibition) varied largely between drugs, ranging larger than 3 x 10(5) ng/mL for 5-FU, larger than 1.5 x 10(2) ng/mL for MMC, 20 ng/mL to 1.4 x 10(3) ng/mL for ADM, and 80 ng/mL to 2.4 x 10(3) ng/mL for CDDP. D0 (dose of radiation reducing the surviving fraction to 37%) ranged from 3.2 to 8.3 Gy (mean +/- standard deviation; 5.8 +/- 1.6 Gy). For hyperthermia, the mean T50 (duration of hyperthermia reducing the surviving fraction to 50%) at 43 degrees C was 9.4 +/- 3.3 min 4.8 to 14.2 min). The T/C ratio at 43 degrees C for 12 min was less than that at 41 degrees C for 30 min (p = .01; the Wilcoxon signed-ranks test). No clear relationship among chemosensitivity, radiosensitivity, hyperthermic sensitivity and pathologic features could be established. CONCLUSIONS: Nine human pancreatic adenocarcinomas varied widely in their sensitivity to chemotherapies, especially for 5-FU. These results suggested that MTT assay may be useful in excluding some less sensitive cases of pancreatic cancer. For hyperthermia, sufficient therapeutic time and temperature may realize enough effect against pancreatic adenocarcinoma.     

Respiratory activity of isolated rat hepatocytes following cold storage and subsequent rewarming: a comparison of sucrose-based and University of Wisconsin solutions

Investigations were carried out on the respiratory function of isolated rat hepatocytes after cold storage alone for periods up to 48 h in either sucrose-based solution (SBS) or University of Wisconsin (UW) solution and after subsequent normothermic preincubation. In both SBS and UW, cold storage for 24 h depressed respiratory function (to 21 +/- 3 and 23 +/- 3 nmol O(2)/min/10(6) cells, respectively) compared to control cell values (31 +/- 3 and 33 +/- 5 nmol O(2)/min/10(6) cells; P < 0.01 in each case). However, normothermic preincubation for 60 min returned respiratory activity to control values (for SBS and UW storage: 41 +/- 6 and 40 +/- 5 nmol O(2)/min/10(6) cells; for control cells: 43 +/- 5 and 46 +/- 6 nmol O(2)/min/10(6) cells). Storage for 48 h in both SBS and UW allowed further depression of respiratory activity, with no recovery after preincubation. Stimulation of respiration by succinate in hepatocytes stored for longer periods was suggestive of increased membrane permeability. Both SBS and UW are effective storage solutions for isolated hepatocytes for up to 24 h as judged by aerobic metabolism, but significant damage was expressed in both solutions when preservation was extended.