Endocrine disruption: thyroid dysfunction in mummichogs (Fundulus heteroclitus) from a polluted habitat.

Mummichogs (Fundulus heteroclitus) from Piles Creeks (PC), New Jersey (a polluted site), are sluggish and show poorer prey capture and predator avoidance than reference fish from Tuckerton (TK). The behavioral dysfunction of the PC fish may be associated with thyroid impairment due to endocrine disruption. In this study, we compared thyroid histology and thyroid hormones in the two populations and determined experimentally whether the polluted environment could alter thyroid hormone levels. PC fish had larger thyroid follicles, greater follicle cell heights, and contained higher plasma thyroxine (T4) levels than TK fish. However, there were no significant differences in either plasma or tissue triiodothyronine (T3). TK fish held in simulated PC environments had higher plasma T4 and lower plasma T3 than field-sampled fish. PC fish held in clean water had lower plasma T4 and T3 than field-sampled fish. In either case, there was no significant difference in tissue T3 content. The contaminants in PC alter thyroid structure and function, which may relate to the behavioral differences between fish from the polluted and reference sites.

Seasonal specificity of hormonal, behavioral, and coloration responses to within- and between-sex encounters in male lizards (Sceloporus undulatus).

This study reports the gender and seasonal specificity of hormonal, behavioral, and coloration responses displayed by "resident" male lizards (Sceloporus undulatus) exposed to male or female "intruders" during staged encounters in outdoor enclosures. Resident males were engaged in staged encounters with males or females for 1 h per day on 9 consecutive days during the breeding and postbreeding seasons. Male-specific responses occurred during the breeding but not the postbreeding season. These included (1) a transient increase in plasma testosterone (T) that was evident on Day 4 and had subsided by Day 10, (2) behavioral displays of aggression (full shows and chases), and (3) a lightening of dorsal integumental color. Female-specific behavioral responses (nod sets) were displayed in both seasons. Season-specific responses consisted only of a transient increase in plasma corticosterone (B) during the breeding season that was evident on Day 4 and had subsided by Day 10. Pushups were displayed in response to both genders during both seasons, although the frequency of pushups was significantly higher in response to females than to males during the postbreeding season. The coloration of residents did not change in response to male intruders during the postbreeding season or to females during either season. These results define the gender and seasonal specificity of hormonal, behavioral, and coloration responses of resident male S. undulatus in social interactions with conspecifics. Thus, our results clarify the biological significance of these responses in terms of potentially aggressive versus courtship interactions and breeding versus postbreeding contexts.

Jumping performance of hylid frogs measured with high-speed cine film.

Jumping performance at 20 degrees C was assessed in five species of hylid frogs using high-speed cine film. Mean takeoff velocities (Vt) varied from 1.5 to 2.4 ms-1 among the species. Peak Vt varied from 1.9 to 2.9 ms-1. Body-mass-specific power output averaged over the entire takeoff period varied from 29 to 91 W kg-1 during the jumps with the highest takeoff velocities. These values are similar to those predicted from jumping distance. As the mass of muscles available to power the jump probably amounts to no more than 17% of the body mass, average muscle-mass-specific power can be over 500 W kg-1. The performance during jumping is even more impressive in view of the fact that the peak power during takeoff is about twice the average power. These frogs must use elastic storage to redistribute power during takeoff to produce the peak power required and may use pre-storage of elastic energy to boost the average power available.

Growth and behavior of thyroid-deficient lizards (Sceloporus undulatus).

This study investigates thyroid control of growth and energy metabolism plus growth-dependent and growth-independent behavioral effects of thyroid manipulation in lizards. Experiments were done on surgically thyroidectomized (Tx) and sham-operated (Sh) yearling Sceloporus undulatus enclosed in their natural habitat. Lizards were placed in an outdoor enclosure in early August. Growth rate was measured and behavior was observed until mid-October. Subsequently, lizards were returned to the lab for measurements of plasma thyroxine (T4), standard metabolic rate (SMR), and running endurance. Thyroidectomy reduced plasma T4 from 4.3 +/- 0.56 ng/ml to undetectable levels (P = 0.006) and SMR by 44% (P less than 0.0001). Thyroid deficiency produced a threefold reduction in growth rate (Tx: 0.04 +/- 0.010 mm/day, n = 12 vs Sh: 0.11 +/- 0.006 mm/day, n = 11, P less than 0.0001). Growth rate was correlated with SMR among individuals (length-specific: r = 0.55, P = 0.027, n = 16), even after statistical removal of mass and treatment effects. The total volume of oxygen consumed for standard metabolism during the growth period, as estimated from allometric equations, was correlated with cumulative growth (R2 = 0.94, P = 0.013) and was significantly lower for thyroid-deficient lizards than for controls (P less than 0.0001). Distance moved in the field and running endurance on a treadmill both scaled on body mass (M0.98 +/- 0.418, P = 0.030 and M1.72 +/- 0.763, P = 0.040, respectively), indicating that larger lizards moved farther and had greater stamina than their smaller counterparts. Neither of these behavioral factors was affected directly by thyroid status. Thyroid-deficient lizards were active for a smaller percentage of the day than controls (Tx: 42.6 +/- 5.7%, n = 11 vs Sh: 57.4 +/- 3.2%, n = 12, P = 0.040), independent of differences in body size. On an interindividual basis, the frequency of activity was significantly correlated with SMR (r = 0.57, P = 0.020, n = 16) and growth rate (mass-specific: r = 0.47, P = 0.025, n = 23).

Interactive effects of thyroxine and experimental location on running endurance, tissue masses, and enzyme activities in captive versus field-active lizards (Sceloporus undulatus).

This study investigates the effects of exogenous thyroxine (T4) on running endurance, tissue masses, and the activities of citrate synthase (CS), pyruvate kinase (PK), cytosolic alpha-glycerophosphate dehydrogenase (alpha-GPDH), and beta-hydroxyacyl Coenzyme A dehydrogenase (HOAD) in Sceloporus undulatus (eastern fence lizard). The enzymes were assayed to indicate maximal catabolic activities that support exercise. Parallel experiments were done on captive and field-active groups to determine whether responses in captive studies adequately predict responses in nature. Exogenous T4 was administered via intraperitoneal pellets. The effect of T4 on running endurance was dependent on the location of the experiment (P = 0.040) such that stamina was increased by T4 only in field-active lizards. At lower levels of biological organization, interactivity between T4 and experimental location was evident but less prevalent than at the level of the whole animal, and some location effects occurred independent of T4 treatment. Heart and kidney masses were significantly greater and total hind leg muscle mass was less in captive than in field-active lizards. Thyroxine reduced liver mass in both locations and kidney mass only in captive lizards. Mass-specific CS and alpha-GPDH in gastrocnemius muscle (mixed fiber type) and HOAD in heart were lower in captive than in field-active lizards; PK in heart and liver and alpha-GPDH in heart were higher in captive lizards. Thyroxine increased CS in liver and HOAD in heart, decreased alpha-GPDH in liver in both locations, and decreased alpha-GPDH in gastrocnemius only in captive lizards. The effects of T4 differed significantly between experimental locations in gastrocnemius muscle (T4 decreased PK only in captive lizards) and in liver (T4 increased PK in field-active lizards and decreased PK in captive lizards). The mechanistic basis of differences in stamina between captive and field-active and between placebo and T4-treated lizards is largely unexplained by the factors measured here, thus illustrating the uncertainty of predicting organismal performance from lower level measurements. Nonetheless, T4 has now been shown to have greater physiological activity in field-active than in captive Sceloporus with regard to resting and total daily metabolic rates and running endurance. The results of this study further confirm that endocrine experiments on captive animals may not predict responses in nature. Further efforts to clarify the physiological significance of seasonal variations in levels of thyroid hormones will have to involve, at least in part, invasive studies on field-active lizards.

Thyroid regulation of resting metabolic rate and intermediary metabolic enzymes in a lizard (Sceloporus occidentalis).

This study investigates the effects of physiological increments in plasma thyroxine (T4) at three levels of biological organization in thyroid-intact and thyroidectomized captive western fence lizards, Sceloporus occidentalis. Two doses of T4-loaded pellets elevated plasma T4 in thyroid-intact lizards from 4.8 +/- 0.47 to 10.7 +/- 2.25 and 20.4 +/- 5.77 ng/ml (mean +/- SE). Surgical thyroidectomy reduced T4 to 1.8 +/- 0.23 ng/ml, and subsequent T4 pellet implantation raised T4 to 14.8 +/- 4.30 ng/ml. Minimal resting metabolic rate (= standard metabolic rate; SMR), a common organismal metric of thyroid perturbation, was reduced 31% (P less than 0.0001) by thyroidectomy and was restored by T4 replacement but was not stimulated by T4 supplementation in thyroid-intact lizards. In T4-replaced, thyroidectomized lizards, SMR was significantly correlated with plasma T4 (r2 = 0.626, P = 0.003, n = 11). At the organ level, liver mass was not changed by any treatment; heart mass was decreased by thyroid deficiency and restored by T4 replacement. At the molecular level, citrate synthase activity was significantly reduced by thyroidectomy and was returned to control levels by T4 replacement in liver and skeletal muscle (gastrocnemius) but was not changed in cardiac muscle. Citrate synthase was not affected in any tissue by T4 supplementation in thyroid-intact lizards. Pyruvate kinase activity was not affected by any of the treatments in any of the tissues. Cytosolic alpha-glycerophosphate dehydrogenase was significantly reduced in liver by all treatments and in skeletal muscle by T4 replacement after thyroidectomy. These results indicate that SMR and cardiac muscle mass in lizards are dependent on normal thyroid function and are expressed maximally in euthyroid animals. The stimulatory effect of T4 on SMR in thyroid-intact lizards, which has been reported previously by several investigators, is a nonphysiological response to pharmacological T4 levels, at least in these captive lizards. Molecular responses are tissue and enzyme dependent and cannot be generalized. Pellet implantation is an effective means of inducing physiological increments in plasma T4 and should replace previously used injection protocols. This new method can be used in capture-recapture experiments involving field-active lizards.

De novo synthesis of adenine nucleotides in different skeletal muscle fiber types.

Management of adenine nucleotide catabolism differs among skeletal muscle fiber types. This study evaluated whether there are corresponding differences in the rates of de novo synthesis of adenine nucleotide among fiber type sections of skeletal muscle using an isolated perfused rat hindquarter preparation. Label incorporation into adenine nucleotides from the [1-14C]glycine precursor was determined and used to calculate synthesis rates based on the intracellular glycine specific radioactivity. Results show that intracellular glycine is closely related to the direct precursor pool. Rates of de novo synthesis were highest in fast-twitch red muscle (57.0 +/- 4.0, 58.2 +/- 4.4 nmol.h-1.g-1; deep red gastrocnemius and vastus lateralis), relatively high in slow-twitch red muscle (47.0 +/- 3.1; soleus), and low in fast-twitch white muscle (26.1 +/- 2.0 and 21.6 +/- 2.3; superficial white gastrocnemius and vastus lateralis). Rates for four mixed muscles were intermediate, ranging between 32.3 and 37.3. Specific de novo synthesis rates exhibited a strong correlation (r = 0.986) with muscle section citrate synthase activity. Turnover rates (de novo synthesis rate/adenine nucleotide pool size) were highest in high oxidative muscle (0.82-1.06%/h), lowest in low oxidative muscle (0.30-0.35%/h), and intermediate in mixed muscle (0.44-0.55%/h). Our results demonstrate that differences in adenine nucleotide management among fiber types extends to the process of de novo adenine nucleotide synthesis.

Thermal adaptations in lizard muscle function.

This study was undertaken to investigate thermal adaptations in muscle contractile properties in closely-related lizards with different preferred body temperatures (PBT). The species examined all belong to the Sphenomorphus group of Australian skinks (Scincidae: Lygosominae). Preferred body temperatures are conservative at the generic level as follows: Ctenotus, 35 degrees C; Sphenomorphus, 30 degrees C; Eremiascincus, 25 degrees C. Contractile properties of the fast glycolytic portion of the iliofibularis muscle were measured. Translational adaptations are evident in several isometric factors, including tetanic tension (Po), twitch tension (Pt), twitch time to peak tension (TPT), and twitch half-relaxation time (1/2 RT). Capacity adaptations are not evident in rates of tetanic tension development (dPo/dt) or in maximal velocities of isotonic shortening (Vmax). Rotational adaptations are not evident in any contractile properties. Thermal limits on upper response temperatures are about 5 degrees C warmer in Ctenotus than in the more cryophilic species, indicative of resistance adaptation in muscle performance. Despite these adaptive shifts, there is little indication that muscle functional capacities are optimized or equalized at PBT in these lizards.

Reduced running endurance in gluconeogenesis-inhibited rats.

The functional significance of gluconeogenesis in prolonging endurance during submaximal activity was assessed in untrained and endurance-trained rats. Gluconeogenesis was inhibited at the phosphoenolpyruvate carboxykinase reaction by 3-mercaptopicolinic acid (3-MPA). Endurance was significantly reduced by 3-MPA in untrained (-32%; P less than 0.005) and in trained rats (-26%; P less than 0.001). Metabolic correlates of fatigue were examined in trained rats. At exhaustion, 3-MPA-treated rats had only 3% of resting hepatic glycogen, 46% of resting white quadriceps glycogen, and 37% of resting blood glucose. All of these substrates were at higher levels in sham-injected controls after the same duration of running (130 min). Glycogen levels in red quadriceps, blood lactate levels, and blood glycerol levels were not different between groups. Plasma free fatty acid levels were elevated to the same extent in both groups after 90 min of activity, remained high at 130 min in controls, but had returned to resting levels in the severely hypoglycemic 3-MPA-treated rats at exhaustion. The results indicate that gluconeogenesis is important for maintaining blood glucose levels and for prolonging endurance time during submaximal activity.

Reduced aerobic capacity and locomotory endurance in thyroid-deficient lizards.

This study was undertaken to examine the effects of thyroid hormonal deficiency on (1) standard (SMR) and maximal (VO2max) rates of O2 consumption, (2) tissue glycolytic and oxidative capacities and (3) submaximal locomotory endurance in a lizard (Dipsosaurus dorsalis). Surgical thyroidectomy induced hypothyroidism in all animals as determined by levels of plasma thyroxine. Hypothyroid lizards had lower levels of SMR (-48%), VO2max (-16%) and citrate synthase activity in liver, heart and skeletal muscle compared to controls. There was a correlated decrease in locomotory endurance in thyroid-deficient animals. Pyruvate kinase activity (an index of glycolytic capacity) in all tissues, and myofibrillar ATPase activity (an index of contractile velocity) in white iliofibularis muscle, showed no significant changes in thyroid-deficient animals. Thyroid hormones appear to be important in ultimately establishing an animal's capacity for locomotory endurance. These findings suggest a new selective context for understanding the evolution of thyroid function.

The effect of body temperature on the locomotory energetics of lizards.

Oxygen consumption (VO2), carbon dioxide production (VCO2), and stamina were measured in the lizard Tupinambis nigropunctatus running at sustainable and non-sustainable velocities (v) on a motor-driven treadmill. Three experimental groups were measured: field-fresh animals at body temperature (Tb) = 35 degrees C and laboratory-maintained animals at Tb = 35 and 25 degrees C. Mean preferred Tb was determined to be 35.2 degrees C. At 35 degrees C, field-fresh animals had a greater maximal oxygen consumption (VO2max corr) (4.22 vs 3.60 ml O2 g-0.76h-1) and a greater endurance. The net cost of transport (slope of VO2 on v) did not differ between the groups (= 2.60 ml O2 g-0.76)km-1). Velocity at which VO2max is attained (MAS) is 0.84 km h-1. The respiratory exchange ratio (R) exceeded 1.0 at v above MAS, indicating supplementary anaerobic metabolism. At 25 degrees C, VO2max corr was lower (2.34 ml O2 g-0.76h-1) as was endurance, MAS occurring at 0.5 km h-1. Net cost of transport was not significantly different than at 35 degrees C. The effect of Tb on locomotory costs was analyzed for this lizard and other species. It was concluded that the net cost of transport is temperature independent in all species examined and the total cost of locomotion (VO2 v-1) is temperature dependent in Tupinambis (Q10 = 1.4-2.0) and all other species examined except one. The energetic cost of locomotion [(VO2active-VO2rest)v-1], previously reported to be temperature independent in lizards, is temperature dependent in Tupinambis (Q10 = 1.3-1.6) and in two other species.2r

Effects of thyroxine supplementation on metabolic rate and aerobic capacity in a lizard.

Effects of thyroxine (T4) on standard and maximal rates of O2 consumption and on citrate synthase activity, an index of mitochondrial oxidative capacity, were examined in the lizard Dipsosaurus dorsalis. Two weeks of daily injections of 200 ng T4/g body mass resulted in significant increases in standard (+60%) and maximal (+15%) rates of O2 consumption. Citrate synthase activity in liver and in red iliofibularis and gastrocnemius muscles was significantly higher than in controls. Correlational analyses indicate that intraspecific differences in standard metabolic rate may be partially attributable to fundamental differences in tissue metabolic capacities and that thyroxine injections lead to a general increase in aerobic capacities. Similar increases in aerobic capacities may occur in response to high thyroid activity during Spring in temperate-zone lizards. The injection protocol used in this study, taken from previous authors, results in suppression of natural thyroid secretory activity and in a 24-h average plasma T4 level of 95 ng/ml compared with 2.5 ng/ml in controls.

Thermal dependence of endurance and locomotory energetics in a lizard.

The thermal dependencies of endurance and the rates of oxygen cnsumption (VO2) and carbon dioxide production (VCO2) were determined for the lizard Dipsosaurus dorsalis walking on a treadmill. The thermal dependencies of endurance and maximal VO2 (VO2 max) are nerly identical. The maximal sustainable speed and the speed at which VO2 max is attained at each experimental body temperature are the same. The net cost of locomotion (the slope of VO2 vs. speed) is termperature independent. Resting VO2 and the incremental change in VO2 required to support locomotion at a sustainable speed are higher at 40 degrees C than at 25 degrees C. Therefore, the total cost of locomotion (VO2 divided by speed) is greater at 40 degrees C. However, the minimum cost of transport, attained at the maximal sustainable speed, is temperature independent. The results clearly indicate that maintenance of a high body temperature is energetically expensive for Dipsosaurus, but that the associated expanded activity capacity enables this lizard to avoid an obligate increase in the energetic requirements of locomotion.