Cellular signalling of PCH-induced pigment aggregation in the crustacean Macrobrachium potiuna erythrophores

The cellular system responsible for the transduction of the pigment-concentrating hormone (PCH) signal was investigated in erythrophores of the freshwater shrimp, Macrobrachium potiuna. Dose-response curves to the hormone were determined in the absence and in the presence of several drugs that affect sequential steps of the Ca²⁺-dependent signalling pathway. Additionally, the ability of forskolin to induce pigment dispersion was evaluated. Neomycin sulphate (10⁻⁴ and 10⁻³ mol · l⁻¹), trifluoperazine (10⁻⁵ and 10⁻⁴ mol · l⁻¹), 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (10⁻⁷ and 10⁻⁵ mol · l⁻¹) and okadaic acid (10⁻⁷ mol · l⁻¹) significantly (P<0.05) decreased the responses to PCH. However, okadaic acid at low concentration (10⁻⁹ mol · l⁻¹) and cyclosporin A (10⁻⁶ and 10⁻⁵ mol · l⁻¹) did not significantly (P>0.05) affect PCH activity. Forskolin (10⁻⁴ mol · l⁻¹) was able to half-maximally reverse the hormone-induced aggregation. Our results suggest that the pigment-concentrating hormone induces pigment aggregation through a Ca²⁺-dependent pathway with a posteriori phosphatase activation, probably the serine/threonine phosphatase 1. Accepted: 30 June 1997     

Coding of blend ratios of binary mixtures by olfactory neurons in the Florida spiny lobster, Panulirus argus

The aim of this study was to investigate quality coding of blend ratios of binary mixtures by olfactory receptor cells in the spiny lobster. Three odorants (adenosine-5′-monophosphate, l-glutamate, and taurine) at 0.1–100 μmol · l⁻¹ and seven blend ratios of each of their binary mixtures at a total concentration of 100 μmol · l⁻¹ were used. The olfactory cells recorded (n = 48) evoked across-neuron patterns for single odorants that were well separated from each other. Across-neuron patterns varied with stimulus concentration but less than with stimulus type. Blend ratios of the three mixtures evoked across-neuron patterns that were orderly placed within a continuum between those elicited by the components. Mixture interactions, defined as a lack of independent effects by a mixture's components, occurred in 25, 24 and 37% of responses to blend ratios of glutamate/taurine, adenosine-5′-monophosphate/taurine, and glutamate/adenosine-5′-monophosphate, respectively. These mixture interactions did not have a large enough effect on the across-neuron patterns for the mixtures such they would be novel relative to those of the single components. These results suggest that despite mixture interactions the quality of individual compounds is not lost when mixed. This corroborates behavioral studies showing that spiny lobsters have the ability to elementally process odor mixtures. Accepted: 23 August 1996     

Protective effects of prostaglandins in the isolated gastric mucosa of the eel, Anguilla anguilla

The protective effect of endogenous prostaglandins on the fish gastric mucosa was evaluated by studying the effect of indomethacin and aspirin, known cyclooxigenase inhibitors, on the mucosal ulceration in the isolated gastric sacs of Anguilla anguilla. Gastric sacs devoid of muscle layers were incubated in the presence of indomethacin (10⁻⁴ mol · l⁻¹) or aspirin (10⁻⁴ mol · l⁻¹) in different experimental conditions. Both the anti-inflammatory drugs produced ulcers, but the effects were more severe in the presence of histamine and in the absence of HCO₃ ⁻ in the incubation bath. The effects of prostaglandin E₂ (PGE₂) on acid secretion rate (J_H) and on alkaline secretion rate (J_OH) were evaluated (with the aid of the pH stat method) in isolated gastric mucosa mounted in Ussing chambers. We found that PGE₂ (10⁻⁸–10⁻⁵ mol · l⁻¹) increased J_H in a dose-dependent manner. In tissues pretreated with luminal omeprazole (10⁻⁴ mol · l⁻¹), PGE₂ stimulated gastric alkaline secretion. It was nullified by serosal removal of HCO₃ ⁻ or Na⁺ and by serosal ouabain (10⁻⁴ mol · l⁻¹). These results suggested that prostaglandins also exert their protective effects in fish gastric mucosa. This protection seems partially due to a stimulation of exogenous HCO₃ ⁻ transport from the serosal to the mucosal side. It is likely that this transport is an active transcellular mechanism coupled to Na⁺ transport. Accepted: 14 April 2000     

The effect of endurance exercise at moderate altitude on serum lipid peroxidation and antioxidative functions in humans

We investigated the effect of training and racing at moderate altitude (MA) on oxidative stress by assessment of serum diene conjugation (DC) and serum antioxidant potential (TRAP). Nine male top level skiers were studied during a national race (20–30 km) at sea level (SL). Thereafter, the athletes trained for 2 weeks at MA, after which they participated in a 20–30 km race at MA. Venous blood samples were taken before and after the race. The DC, indicating early events of lipid peroxi dation, did not change during the race at SL (16 850 vs 15 900 ΔAbsorbance · l⁻¹) or at MA (19 870 vs. 20 630 ΔAbs · l⁻¹). At MA serum DC was higher than at SL both before (25%) and after (30%) the race, the postrace difference being statistically significant (P < 0.05). The TRAP increased during the race at MA (from 1387 to 1943 μmol · 1⁻¹, P  =  0.016), but not at SL (1713 vs 1582 μmol · l⁻¹). These observations would suggest that the level of oxidative stress might be greater during living, training and racing at MA (higher DC levels). Increased TRAP during the race at MA may indicate that the physiological adaptation to extreme acute oxidative stress was altered. The physiological significance of this observation remains to be investigated. Accepted: 18 October 1996     

Insulin and glucagon immunoreactivity during high-intensity exercise under opiate blockade

Eight fit men [maximum oxygen consumption (O_2max) 64.6 (1.9) ml · kg⁻¹ · min⁻¹, aged 28.3 (1.7) years (SE in parentheses) were studied during two treadmill exercise trials to determine the effect of endogenous opioids on insulin and glucagon immunoreactivity during intense exercise (80% O_2max). A double-blind experimental design was used with subjects undertaking the two exercise trials in counterbalanced order. Exercise trials were 20 min in duration and were conducted 7 days apart. One exercise trial was undertaken following administration of naloxone (N; 1.2 mg; 3 ml) and the other after receiving a placebo (P; 0.9% NaCl saline; 3 ml). Prior to each experimental trial a flexible catheter was placed into an antecubital vein and baseline blood samples were collected. Immediately after, each subject received either a N or P bolus injection. Blood samples were also collected after 20 min of continuous exercise (running). Glucagon was higher (P < 0.05), while insulin was lower (P < 0.05), during exercise compared with pre-exercise values in both trials. However, glucagon was higher (P < 0.05) in the P than in the N exercise trial [141.4 (8.3) ng · l⁻¹ vs 127.2 (7.6) ng · l⁻¹]. There were no differences in insulin during exercise between the P and N trials [50.2 (4.3) pmol · l⁻¹ vs 43.8 (5) pmol · l⁻¹]. These data suggest that endogenous opioids may augment the glucagon response during intense exercise. Accepted: 15 June 1996     

Evidence for NO-dependent vasodilation in the trout (Oncorhynchus mykiss ) coronary system

The effects of l-arginine, and its analogues N ^ω-nitro-l-arginine methyl ester and N ^ω-nitro-l-arginine on vascular resistance were investigated in the intact coronary system of an isolated non-working trout heart preparation. l-Arginine, at 10^–8 mol · l^–1induced a slight vasodilatory effect (max 10%). N ^ω-nitro-l-arginine methyl ester and N ^ω-Nitro-l-arginine in the range 10^–8–10^–4 mol · l^–1 caused dose-dependent increases in coronary resistance. The vasodilatory action of l-arginine was abolished when the preparation was pretreated with 10^–4 mol · l^–1 N ^ω-nitro-l-arginine or N ^ω-nitro-l-arginine methyl ester. Nitroprusside alone at 1 mmol · l^–1 induced a maximum vasodilation (30%) of the coronary system. Methylene blue a known inhibitor of guanylate cyclase, induced a strong vasoconstriction (already significant at 10^–5 mol · l^–1) and was able to overcome the vasodilative effect of nitroprusside. The endothelial nitric oxide agonists acetylcholine and serotonin, established in mammalian vessels, also mediate vasodilation in trout coronary system. In 50% of preparations, acetylcholine induced a biphasic response with vasodilation at low concentration (max 15% at 10^–8 mol · l^–1). Serotonin displayed a dose-response vasodilation in the range 10^–8–10^–4 mol · l^–1 (max 20%). These vasodilative effects were reduced or abolished by 10^–4 mol · l^–1 l-NA. These data support the existence of NO-mediated vasodilation mechanisms in the trout coronary system. Accepted: 1 July 1996     

Differences among plant species in cuticular permeabilities and solute mobilities are not caused by differential size selectivities

Solute mobilities in cuticular membranes of six species (Hedera helix, Malus domestica, Populus alba, Pyrus communis, Stephanotis floribunda, Strophantus gratus) were measured using plant hormones, growth regulators and other organic model compounds varying in molar volumes from 99 to 349 mL · mol⁻¹ The dependence of mobilities (k*) on molar volume (V _x ) was exponential and could be described with equations of the type log k*=log k*⁰−^′ V _x . The y-intercepts (log k*⁰) represent mobilities of a hypothetical solute of zero molar volume. The parameter β′ is a measure of size selectivity of cuticular membranes and no differences among the six species were observed. At 25 °C the average β′ was 0.0095 mol · mL⁻¹. Solute mobility decreased by about a factor of 8.9 when molar volume increased by 100 mL · mol⁻¹ and the mobility of a compound with V _x  = 100 mL · mol⁻¹ was about 700-fold higher than the mobility of a compound with V _x  = 400 mL · mol⁻¹. Size selectivity decreased with increasing temperatures and for Strophantusβ′-values of 1.6 × 10⁻² to 8.0 × 10^-4 mol · mL⁻¹ were obtained for 10 and 30 °C, respectively. The-intercepts (log k*⁰) differed among plant species by 3 orders of magnitude and since size selectivity was the same for all species, solute mobilities for solutes having zero molar volumes were the sole cause for differences among species in solute mobilities and permeabilities. We argue that these differences in k*⁰ are related to tortuosity of the diffusion path. These results were used to derive an equation which predicts rates of cuticular penetration on the basis of k*⁰, the average size selectivity of 9.5 × 10⁻³ mol · mL⁻¹ and the driving forces of penetration. Received: 25 November 1997 / Accepted: 9 March 1998     

Effect of circulatory occlusion on human muscle metabolism during exercise and recovery

To assess muscle metabolism and inorganic phosphate (P_i) peak splitting during exercise, 31-phosphorus nuclear magnetic resonance spectroscopy was performed during ramp incremental and submaximal step exercise with and without circulatory occlusion. Seven healthy men performed calf flexion in a superconducting magnet. There was no P_i splitting during ramp incremental exercise with the circulation present and phosphocreatine (PCr) decreased linearly by 0.07 (SEM 0.01) mmol · l⁻¹ · s⁻¹, while exercise with the circulation occluded caused the P_i peak to split into a high and a low pH peak. The rate of PCr decrease during exercise with the circulation occluded was 0.15 (SEM 0.03) mmol · l⁻¹ · s⁻¹ which with the efficiency of the adenosine 5′-triphosphate (ATP) hydrolysis reaction corresponded well to the mechanical energy. Both with and without occlusion of the circulation PCr decreased with some time lag which may reflect the consumption of residual oxygen. In submaximal step exercise PCr decreased exponentially at the onset of exercise with the circulation open whereas it decreased linearly by 0.15␣mmol · l⁻¹ · s⁻¹ when the circulation was occluded. After exercise, occlusion of the circulation was maintained for 1 min more and there was no PCr resynthesis. It is suggested that ATP synthesis was limited by the availability of oxygen. Accepted: 14 August 1996     

Active calcium ion transport in Xenopuslaevis skin

The skin of intact, free-swimming Xenopus laevis transports Ca²⁺ inwardly in a manner that is proportional to the external [Ca²⁺] up to about 0.3 mmol · l⁻¹, saturates above 0.3 mmol · l⁻¹, and is opposed to the electrochemical gradient. Efflux is relatively constant at external concentrations between 0.016 and 0.6 mmol · l⁻¹; net flux which is negative below 0.125 mmol · l⁻¹ becomes positive above this external [Ca²⁺]. Allometric analysis suggests that both Ca²⁺ influx and efflux scale to the 2/3 power approximately like surface area. There were no significant differences in influx between summer and fall animals; however, efflux was greater in the fall and this resulted in a change from positive balance in the summer to negative balance in the fall. Isolated skins were shown to support a Ca²⁺ uptake rate of nearly 30 nmol · cm⁻² · h⁻¹. The phenylalkylamine verapamil in the apical bathing solution significantly inhibited this at 25 μmol · l⁻¹. The benzothiazepine diltiazem was also effective at 50 μmol · l⁻¹ while the dihydropyradine nifedipine was ineffective up to 100 μmol · l⁻¹. The inorganic ion La³⁺ was effective at blocking Ca²⁺ uptake at 300 μmol · l⁻¹; Ni²⁺ was also effective at 500 μmol · l⁻¹ but Co²⁺ was ineffective up to 500 μmol · l⁻¹. These results suggest that apical calcium channels in Xenopuslaevis skin have properties similar to mammalian L-channels and fish gill Ca²⁺ channels. Accepted: 23 January 1997     

Active excretion of ammonia across the gills of the shore crab Carcinus maenas and its relation to osmoregulatory ion uptake

The mechanism of transbranchial excretion of total ammonia of brackish-water acclimated shore crabs, Carcinus maenas was examined using isolated, perfused gills. Applying physiological gradients of NH₄Cl (100–200 μmol · l⁻¹) directed from the haemolymph space to the bath showed that the efflux of total ammonia consisted of two components. The saturable component (excretion of NH₄ ⁺) greatly exceeded the linear component (diffusion of NH₃). When an outwardly directed gradient (200 μmol · l⁻¹) was applied, total ammonia in the perfusate was reduced by more than 50% during a single passage of saline through the gill. Effluxes of ammonia along the gradient were sensitive to basolateral dinitrophenol, ouabain, and Cs⁺ and to apical amiloride. Acetazolamide (1 mmol · l⁻¹ basolateral) or Cl⁻-free conditions had no substantial effects on ammonia flux, which was thus independent of both carbonic anhydrase mediated pH regulation and osmoregulatory NaCl uptake. When an inwardly directed gradient (200 μmol · l⁻¹) was employed, influx rates were about 10-fold smaller and unaffected by basolateral ouabain (5 mmol · l⁻¹) or dinitrophenol (0.5 mmol · l⁻¹). Under symmetrical conditions (100 μmol · l⁻¹ NH₄Cl on both sides) ammonia was actively excreted against the gradient of total ammonia, which increased strongly during the experiment and against the gradient of the partial pressure of NH₃. The active excretion rate was reduced to 7% of controls by basolateral dinitrophenol (0.5 mmol · l⁻¹), to 44% by basolateral ouabain (5 mmol · l⁻¹), to 46% by Na⁺-free conditions and to 42% by basolateral Cs⁺ (10 mmol · l⁻¹), indicating basolateral membrane transport of NH₄ ⁺ via the Na⁺/K⁺-ATPase and K⁺-channels and a second active, apically located, Na⁺ independent transport mechanism of NH₄ ⁺. Anterior gills, which are less capable of active ion uptake than posterior gills, exhibited even increased rates of active excretion of ammonia. We conclude that, under physiological conditions, branchial excretion of ammonia is a directed process with a high degree of effectiveness. It even allows active extrusion against an inwardly directed gradient, if necessary. Accepted: 11 March 1998     

The Impact of heat exposure and repeated exercise on circulating stress hormones

To determine if heat exposure alters the hormonal responses to moderate, repeated exercise, 11 healthy male subjects [age = 27.1 (3.0) years; maximal oxygen consumption, V˙O_2max = 47.6 (6.2) ml · kg · min⁻¹; mean (SD)] were assigned to four different experimental conditions according to a randomized-block design. While in a thermoneutral (23°C) or heated (40°C, 30% relative humidity) climatic chamber, subjects performed either cycle ergometer exercise (two 30-min bouts at ≈50% V˙O_2max, separated by a 45-min recovery interval, CEx and HEx conditions), or remained seated for 3 h (CS and HS conditions). Blood samples were analyzed for various exercise stress hormones [epinephrine (E), norepinephrine (NE), dopamine, cortisol and human growth hormone (hGH)]. Passive heating did not alter the concentrations of any of these hormones significantly. During both environmental conditions, exercise induced significant (P < 0.001) elevations in plasma E, NE and hGH levels. At 23°C during bout 1: E = 393 (199) pmol · l⁻¹ (CEx) vs 174 (85) pmol · l⁻¹ (CS), NE = 4593 (2640) pmol · l⁻¹ (CEx) vs 1548 (505) pmol · l⁻¹ (CS), and hGH = 274 (340) pmol · l⁻¹ (CEx)vs 64 (112) pmol · l⁻¹ (CS). At 40°C, bout 1: E = 596 (346) pmol · l⁻¹ (HEx) vs 323 (181) pmol · l⁻¹ (HS), NE = 7789 (5129) pmol · l⁻¹ (HEx) vs 1527 (605) pmol · l⁻¹ (HS), and hGH = 453 (494) pmol · l⁻¹ (HEx) vs 172 (355) pmol · l⁻¹ (HS). However, concentrations of plasma cortisol were increased only in response to exercise in the heat [HEx = 364 (168) nmol · l⁻¹ vs HS = 295 (114) nmol · l⁻¹). Compared to exercise at room temperature, plasma levels of E, NE and cortisol were all higher during exercise in the heat (P < 0.001 in all cases). The repetition of exercise did not significantly alter the pattern of change in cortisol or hGH levels in either environmental condition. However, repetition of exercise in the heat increased circulatory and psychological stress, with significantly (P < 0.001) higher plasma concentrations of E and NE. These results indicate a differential response of the various stress hormones to heat exposure and repeated moderate exercise. Accepted: 16 April 1997     

Significant changes in VLDL-Triacylglycerol and glucose tolerance in obese subjects following ten days of training

We characterized the effect of ten days of training on lipid metabolism in 6 [age 37.2 (2.3) years] sedentary, obese [BMI 34.4 (3.0) kg · m⁻²] males with normal glucose tolerance. An oral glucose tolerance test was performed prior to and at the end of the 10 d of training period. The duration of each daily exercise session was 40 min at an intensity equivalent to ˜75% of the age predicted maximum heart rate. Blood measurements were performed after an overnight fast, before and at the end of the 10 d period. Plasma triacylglycerol was significantly (p < 0.05) reduced following exercise training (2.15 ± 0.29 vs. 1.55 ± 0.28 mmol · l⁻¹). Very low density lipoprotein-triacylglycerol was also significantly (p < 0.05) reduced (1.82 ± 0.3 vs. 1.29 ± 0.29 mmol · l⁻¹). No significant changes in high density lipoprotein-cholesterol were observed as a result of training. Following training fasting plasma glucose and fasting plasma insulin were significantly reduced [Glucose: 5.9 (0.2) mmol · l⁻¹ vs. 5.3 (0.22) mmol · l⁻¹ (p < 0.05); Insulin 264.3 (53.8) ρ · mol · l⁻¹ vs. 200.9 (30.1) ρ · mol · l⁻¹, p = 0.05]. The total area under the glucose curve during the OGTT decreased significantly (p < 0.05). These preliminary data suggest that short-term exercise, without concomitant loss of body mass, induces favorable changes in plasma triacylglycerol, and very low density lipoprotein-triacylglycerol and glucose tolerance but has no effect on high density lipoproteincholesterol. Accepted: 7 January 1998     

A neutral carrier-based liquid membrane microelectrode for divalent putrescine cations

A new ion-selective liquid membrane microelectrode, based on the neutral carrier 1,1′-bis(2,3-naphtho-18-crown-6), is described that shows the dependence of EMF on the activity of divalent putrescine cations a _Put, with the linear slope s _Put = 26 ± 3 mV/decade (mean ± SD, N = 18), in the range 10⁻⁴–10⁻¹ M at 25 ± 1 °C. Values of potentiometric putrescine cation selectivity coefficients of logK ^Pot _{Put j} (mean ± SD, N) are obtained by the separate solution method for the ions K⁺ (1.0 ± 0.4, 10), Na⁺ (−1.2 ± 0.4, 8), Ca²⁺ (−2.3 ± 0.5, 10) and Mg²⁺ (−2.5 ± 0.5, 7). The microelectrode can be applied for the direct analysis of the activities of free divalent putrescine cations in the range 5 × 10⁻⁴ to 10⁻¹ M in an extracellular ionic environment. Established analytical methods, e.g. high performance liquid chromatography, determine the total concentration of the derivatives of free and bound putrescine. Received: 20 December 1998 / Revised version: 7 May 1999 / Accepted: 27 May 1999     

Effects of 3 days of carbohydrate supplementation on muscle glycogen content and utilisation during a 1-h cycling performance

This study compared the effects of supplementing the normal diets of six trained cyclists [maximal oxygen uptake O_2max) 4.5 (0.36)l · min⁻¹; values are mean (SD)] with additional carbohydrate (CHO) on muscle glycogen utilisation during a 1-h cycle time-trial (TT). Using a randomised crossover design, subjects consumed either their normal diet (NORM) for 3 days, which consisted of 426 (137) g · day⁻¹ CHO [5.9 (1.4) g · kg⁻¹ body mass (BM)], or additional CHO (SUPP) to increase their intake to 661 (76) g · day⁻¹ [9.3 (0.7) g · kg⁻¹ BM]. The SUPP diet elevated muscle glycogen content from 459 (83) to 565 (62) mmol · kg⁻¹ dry weight (d.w.) (P < 0.05). However, despite the increased pre-exercise muscle glycogen stores, there was no difference in the distance cycled during the TT [40.41 (1.44) vs 40.18 (1.76) km for NORM and SUPP, respectively]. With NORM, muscle glycogen declined from 459 (83) to 175 (64) mmol · kg⁻¹ d.w., whereas with SUPP the corresponding values were 565 (62) and 292 (113) mmol · kg⁻¹ d.w. Accordingly, both muscle glycogen utilisation [277 (64) vs 273 (114) mmol · kg⁻¹ d.w.] and total CHO oxidation [169 (20) vs 165 (30) g · h⁻¹ for NORM and SUPP, respectively] were similar. Neither were there any differences in plasma glucose or lactate concentrations during the two experimental trials. Plasma glucose concentration averaged 5.5 (0.5) and 5.6 (0.6) mmol · l⁻¹, while plasma lactate concentration averaged 4.4 (1.9) and 4.4 (2.3) mmol · l⁻¹ for NORM and SUPP, respectively. The results of this study show that when well-trained subjects increase the CHO content of their diet for 3 days from 6 to 9 g · kg⁻¹ BM there is only a modest increase in muscle glycogen content. Since supplementary CHO did not improve TT performance, we conclude that additional CHO provides no benefit to performance for athletes who compete in intense, continuous events lasting 1 h. Furthermore, the substantial muscle CHO reserves observed at the termination of exercise indicate that whole-muscle glycogen depletion does not determine fatigue at this exercise intensity and duration. Accepted: 25 November 1996     

The relationship between plasma potassium concentration and muscle torque during recovery following intense exercise

The present study investigated the relationship between plasma potassium ion concentration ([K⁺]) and skeletal muscle torque during three different 15-min recovery periods after fatigue induced by four 30-s sprints. Four males and one female completed the multiple sprint exercise on three separate days; recovery was passive, i.e. no cycling exercise (PRec), active cycling at 30% peak oxygen consumption O_2peak (30% Rec) and active cycling at 60% O_2peak (60% Rec). Plasma [K⁺] was measured from blood sampled from an antecubital vein of subjects at rest and at 0, 3, 5, 10 and 15 min into each recovery. Isokinetic leg strength was measured at rest and at 1, 6, 11 and 16 min during each recovery. Following the exhaustive sprints, [K⁺] increased significantly from an average mean (SEM) resting value of 3.81 (0.07) mmol · l⁻¹ to 4.48 (0.19) mmol · l⁻¹ (P < 0.01). In all recovery conditions, plasma [K⁺] returned to resting levels within 3 min following the fourth sprint. However, in the two active recovery conditions plasma [K⁺] increased over the remainder of the recovery periods to 4.36 (0.12) mmol · l⁻¹ in the 30% Rec condition and 4.62 (0.12) mmol · l⁻¹ in the 60% Rec condition, the latter being significantly higher than the former (P < 0.01). The maximum torque measured following the sprints decreased significantly, on average, to 61.1 (8.36)% of peak levels (P < 0.01). After 15 min of recovery, maximum torque was highest in the 30% Rec condition at 92.13 (3.06)% of peak levels (P < 0.01), compared to 85.23 (3.64)% and 85.71 (0.82)% for the PRec and 60% Rec conditions, respectively. In contrast to the significant differences in plasma [K⁺] across all three recovery conditions, muscle torque recovery was significantly different in only the 30% Rec condition. In summary, recovery of peak levels of muscle torque following fatiguing exercise does not appear to follow changes in plasma [K⁺]. Accepted: 18 October 1996     

Leaf photosynthesis, plant growth and nitrogen allocation in rice under different irradiances

The photosynthetic rates and various components of photosynthesis including ribulose-1,5-bisphosphate carboxylase (Rubisco; EC 4.1.1.39), chlorophyll (Chl), cytochrome (Cyt) f, and coupling factor 1 (CF₁) contents, and sucrose-phosphate synthase (SPS; EC 2.4.1.14) activity were examined in young, fully expanded leaves of rice (Oryza sativa L.) grown hydroponically under two irradiances, namely, 1000 and 350 μmol quanta · m⁻² · s⁻¹, at three N concentrations. The light-saturated rate of photosynthesis measured at 1800 μmol · m⁻² · s⁻¹ was almost the same for a given leaf N content irrespective of growth irradiance. Similarly, Rubisco content and SPS activity were not different for the same leaf N content between irradiance treatments. In contrast, Chl content was significantly greater in the plants grown at 350 μmol · m⁻² · s⁻¹, whereas Cyt f and CF₁ contents tended to be slightly smaller. However, these changes were not substantial, as shown by the fact that the light-limited rate of photosynthesis measured at 350 μmol · m⁻² · s⁻¹ was the same or only a little higher in the plants grown at 350 μmol · m⁻² · s⁻¹ and that CO₂-saturated photosynthesis did not differ between irradiance treatments. These results indicate that growth-irradiance-dependent changes in N partitioning in a leaf were far from optimal with respect to N-use efficiency of photosynthesis. In spite of the difference in growth irradiance, the relative growth rate of the whole plant did not differ between the treatments because there was an increase in the leaf area ratio in the low-irradiance-grown plants. This increase was associated with the preferential N-investment in leaf blades and the extremely low accumulation of starch and sucrose in leaf blades and sheaths, allowing a more efficient use of the fixed carbon. Thus, morphogenic responses at the whole-plant level may be more important for plants as an adaptation strategy to light environments than a response of N partitioning at the level of a single leaf. Received: 23 February 1997 / Accepted: 8 May 1997     

Purification and characterization of lanatoside 15′-O-acetylesterase from Digitalis lanata Ehrh.

Lanatoside 15′-O-acetylesterase (LAE) from in-vitro-cultivated cells of Digitalis lanata Ehrh. was isolated and partially sequenced. The enzyme was extracted with citrate buffer from acetone dry powder. It was purified in a two-step chromatographical procedure including Phenyl Sepharose hydrophobic interaction chromatography followed by CM Sepharose cation-exchange chromatography to more than 330 μmol · s⁻¹ · (g protein)⁻¹. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of the purified protein showed a major band at 39 kDa. The protein was identified by correlation of band intensity on SDS-PAGE and enzyme activity of CM Sepharose column fractions. Size-exclusion chromatography on Sephacryl 200 revealed a single activity peak with an apparent molecular mass of about 85 kDa. Electrophoresis under nondenaturating conditions of purified LAE showed only one band with esterase activity. The intensity of this band was correlated with that of the 39-kDa band after SDS-PAGE. About 30% of the protein, including the N-terminus and several fragments obtained by Lys-C protease digestion, was sequenced. A fragment obtained by Lys-C digestion showed partial homology to other hydrolases and apoplasmic proteins. It included the probable location of an active-site histidine. The activity of LAE was high in non-morphogenic D. lanata cell strains selected for high activities in the chemical transformation of cardenolides, but rather low in the proembryogenic masses of the embryogenic cell strain VIII. It increased during the development of somatic embryos. The LAE activity in leaves of D. lanata plants was in the range 4–24 nmol · s⁻¹ · (g protein)⁻¹. Received: 25 March 1997 / Accepted: 3 July 1997     

Oxygen consumption and sulfide detoxification in the lugworm Arenicola marina (L.) at different ambient oxygen partial pressures and sulfide concentrations

The lugworm Arenicola marina is a typical inhabitant of intertidal flats. In its L-shaped burrow the animal is exposed to varying concentrations of O₂ and toxic sulfide depending on the tides. The lugworm is able to detoxify sulfide through its oxidation to thiosulfate. When exposed to declining O₂ tensions Arenicola marina reacted as an oxyconformer. In the presence of 25 μmol · l⁻¹ sulfide the respiration was not affected. In contrast, the lugworm consumed significantly less O₂ at any Po₂ in the presence of 200 μmol · l⁻¹ sulfide. Without sulfide anaerobic metabolism started at a Po₂ of approximatedly 10 kPa. Even at high O₂ tensions animals exposed to sulfide produced significantly more anaerobic metabolites compared with the controls. Accordingly the critical value Pc_M, the ambient Po₂ below which anaerobic metabolism starts, was shifted towards normoxia. Since O₂ supply was sufficient for aerobic metabolism, anaerobiosis was induced by sulfide. An influx of sulfide was observed at 25 as well as at 200 μmol · l⁻¹ sulfide. The main product of sulfide detoxification in the lugworm was thiosulfate. Its synthesis increased with ambient Po₂ and depended on the sulfide concentration. Sulfide and thiosulfate were detected in the coelomic fluid, the blood, and the body wall of Arenicola marina. Only about 2% of the ambient O₂ was used for sulfide detoxification at 25 μmol · l⁻¹ sulfide and about 50% at 200 μmol · l⁻¹ sulfide, respectively. Even at the low sulfide concentration Arenicola marina's capacity to detoxify sulfide was too low to maintain a complete aerobic metabolism. Accepted: 19 February 1997     

Energetics during nursing and early postweaning fasting in hooded seal (Cystophora cristata ) pups from the Gulf of St Lawrence, Canada

In this study we measured growth and milk intake and calculated energy intake and its allocation into metabolism and stored tissue for hooded seal (Cystophora cristata) pups. In addition, we measured mass loss, change in body composition and metabolic rate during the first days of the postweaning fast. The mean body mass of the hooded seal pups (n = 5) at the start of the experiments, when they were new-born, was 24.3 ± 1.3 kg (SD). They gained an average of 5.9 ± 1.1. kg · day⁻¹ of which 19% was water, 76% fat and 5% protein. This corresponds to an average daily energy deposition of 179.8 ± 16.0 MJ. The pups were weaned at an average body mass of 42.5 ± 1.0 kg 3.1 days after the experiment was initiated. During the first days of the postweaning fast the pups lost an average of 1.3 ± 0.5␣kg of body mass daily, of which 56% was water, 16% fat and 28% protein. During the nursing period the average daily water influx for the pups was 124.6 ± 25.8 ml · kg⁻¹. The average CO₂ production during this period was 1.10 ± 0.20 ml · g⁻¹ · h⁻¹, which corresponds to a field metabolic rate of 714 ± 130 kJ ·  kg⁻¹ · day⁻¹, or 5.8 ± 1.1 times the predicted basal metabolic rate according to Kleiber (1975). During the postweaning fast the average daily water influx was reduced to 16.1 ± 6.6 ml · kg⁻¹. The average CO₂ production in␣this period was 0.58 ± 0.17 ml · g⁻¹ · h⁻¹ which corresponds to a field metabolic rate of 375 ± 108 kJ · kg⁻¹ · day⁻¹ or 3.2 ± 0.9 times the predicted basal metabolic rate. Average values for milk composition were 33.5% water, 58.6% fat and 6.2% protein. The pups drank an average of 10.4 ± 1.8␣kg of milk daily, which represents an energy intake of 248.9 ± 39.1 MJ · day⁻¹. The pups were able to store 73.2 ± 7.7% of this energy as body tissue. Accepted: 15 August 1996     

Effect of dephosphorylated 2′,5′-trioligoadenylate on the entry of sodium ions into human neuroblastoma cells

Using a radioisotope technique, we studied the effect of dephosphorylated 2′,5′-trioligoadenylate (2′,5′ ApApA) on the entry of sodium ions into cultured human neuroblastoma cells (IMR 32 strain). Short-term (nearly 1 h) action of 2′,5′ ApApA did not influence the entry of sodium ions through voltage-operated sodium channels in the absence of neurotoxins modulating the characteristics of these channels (toxin of a scorpion, Leiurus quinquestriatus, + veratrine). At the same time, 2′,5′ ApApA weakened in a dose-dependent manner the entry of sodium ions through sodium channels opened upon the action of the above neurotoxins. In cells cultured for 22 h in a medium containing 5 · 10⁻⁶ M 2′,5′ ApApA, the entry of sodium ions in the absence of neurotoxins was 25% greater, while in the presence of neurotoxins it was 24% smaller than that in the control cells. Tetrodotoxin (TTX, 4 · 10⁻⁷ M) blocked completely sodium entry through sodium channels in cells cultured in the absence of 2′,5′ ApApA, while in cells cultured in the presence of this adenylate TTX decreased the entry by 64%. It is hypothesized that long-lasting action of 2′,5′ ApApA results in the appearance of voltage-operated TTX-insensitive sodium channels in the plasma membrane of IMR 32 cells. Our data show that 2′,5′ ApApA is capable of modulating to a considerable extent the functioning of mechanisms controlling transport of sodium ions in cells of human neuroblastoma cells of the IMR 32 strain. Neirofiziologiya/Neurophysiology, Vol. 40, No. 1, pp. 3–8, January–February, 2008.