Peter L. Lutz

Marine Turtles as Sentinels of Ecosystem Health: Is Fibropapillomatosis an Indicator?

Marine turtle fibropapillomatosis (FP) is a disease primarily affecting green turtles (Chelonia mydas) that is characterized by multiple cutaneous masses. In addition, the condition has been confirmed in other species of sea turtles. The disease has a worldwide, circumtropical distribution and has been observed in all major oceans. Although reported since the late 1930s in Florida, it was not until the late 1980s that it reached epizootic proportions in several sea turtle populations. Long-term studies have shown that pelagic turtles recruiting to near shore environments are free of the disease. After exposure to these benthic ecosystems, FP manifests itself with primary growths in the corner of the eyes spreading to other epithelial tissue. One or more herpesviruses, a papillomavirus, and a retrovirus have been found associated with tumors using electron microscopy and molecular techniques; however, the primary etiological agent remains to be isolated and characterized. Field observations support that the prevalence of the disease is associated with heavily polluted coastal areas, areas of high human density, agricultural runoff, and/or biotoxin-producing algae. Marine turtles can serve as excellent sentinels of ecosystem health in these benthic environments. FP can possibly be used as an indicator but correlations with physical and chemical characteristics of water and other factors need to be made. Further research in identifying the etiologic agent and its association with other environmental variables can provide sufficient parameters to measure the health of coastal marine ecosystems, which serve not only as ecotourism spots but also as primary feeding areas for sea turtles.

Anoxia Tolerant Brains

While medical science has struggled to find ways to counteract anoxic brain damage with limited success, evolution has repeatedly solved this problem. The best-studied examples of anoxia-tolerant vertebrates are the crucian carp and some North American Freshwater turtles. These can survive anoxia for days to months, depending of temperature. Both animals successfully fight any major fall in brain ATP levels, but the strategies they use to accomplish this are quite divergent. The anoxic turtle suppresses brain activity to such a degree that it becomes virtually comatose. The underlying mechanisms involve closing down ion conductances and releasing GABA and adenosine. By contrast, the crucian carp remains active in anoxia, although it suppresses selected brain functions, and avoids lactate self-poisoning by producing an exotic anaerobic end-product. These animals provide unique models for studying anoxic survival mechanisms both on a molecular and physiological level.

Release of adenosine and ATP in the brain of the freshwater turtle (Trachemys scripta) during long-term anoxia

Extracellular adenosine and ATP levels were monitored by microdialysis in the striatum of the freshwater turtle Trachemys scripta during long-term N2 respiration. After an initial rise in extracellular adenosine, a second peak of longer duration and higher in intensity, followed. The frequencies of these adenosine cycles varied considerably between individual turtles, such that the shortest time between the peaks was 80 min and the longest was 300 min (mean 151 min). After about 60 min anoxia, there was also a slow increase in extracellular ATP, rising from a normoxic concentration of 1.21 +/- 0.12 to 7.58 +/- 3.70 nmol l(-1) at 240 min anoxia. The results suggest that adenosine may continue to have a protective function in the turtle brain during long-term anoxia and that extracellular ATP might not function as an excitatory neurotransmitter in the anoxic turtle brain.

Time Course of Anoxia-Induced Increase in Cerebral Blood Flow Rate in Turtles: Evidence for a Role of Adenosine

The exceptional ability of the turtle brain to survive prolonged anoxia makes it a unique model for studying anoxic survival mechanisms. We have used epiillumination microscopy to record blood flow rate in venules on the cortical surface of turtles (Trachemys scripta). During anoxia, blood flow rate increased 1.7 times after 45–75 min, whereupon it fell back, reaching preanoxic values after 115 min of anoxia. Topical super-fusion with adenosine (50 μM) during normoxia caused a 3.8-fold increase in flow rate. Superfusing the brain with the adenosine receptor blocker aminophylline (250 μM) totally inhibited the effects of both adenosine and anoxia, while aminophylline had no effect on normoxic flow rate. None of the treatments affected systemic blood pressure. These results indicate an initial adenosine-mediated increase in cerebral blood flow rate during anoxia, probably representing an emergency response before deep metabolic depression sets in.

Vertebrate brains at the pilot light

While the brains of most vertebrates are unable to tolerate more than a few minutes of anoxia, some freshwater turtles (Trachemys and Chrysemys), crucian carp (Carassius carassius) and frogs (Rana pipens and Rana temporaria) can survive anoxia for hours to months. Obviously, anoxia tolerance has evolved separately several times and this is also reflected in the divergent strategies these animals utilize to survive without oxygen. The turtles and crucian carp defend their brain ATP levels and avoid a loss of ion homeostasis by reducing ATP use. In the turtles, the early release of adenosine and the activation of K(ATP) channels, a progressive release of GABA and a drastic reduction in electric activity and ion fluxes send the brain into a comatose like state. The crucian carp displays a more modest depression of ATP use, probably achieved through a moderated release of GABA and adenosine, allowing the animal to maintain physical activity in anoxia. The anoxic frog, on the other hand, seems to rely on mechanisms that greatly retard the anoxia induced fall in ATP levels and loss of ion homeostasis, so that the brain can be saved as long as the anoxia is limited to a few hours. The sequence of events characterizing the anoxic frog brain is similar to that of failing anoxic mammalian brain, although over a greatly extended time frame, allowing the frog to die slowly in anoxia, rather than survive. By contrast the only factor that limits anoxic survival in turtles and crucian carp may be the final depletion of their glycogen reserves.

Anoxia tolerant animals from a neurobiological perspective

This paper discusses the mechanisms for brain anoxia survival seen in crucian carp (Carassius carassius) and a few species of freshwater turtle (Chrysemys and Trachemys species). Comparisons are made with the hypoxic tolerant mammalian neonate brain. In the anoxic tolerant species the basic strategy for anoxia survival appears to be the maintenance of ion gradients, and thereby the avoidance of anoxic depolarization. Important facilitating factors involve having huge glycogen stores, increased blood supply to the brain, the suppression of electrical activity, increased release of inhibitory neuromodulators and neurotransmitters, upregulation of inhibitory neuroreceptors, the down-regulation of excitatory ion conductance and the down-regulation of Ca2+ channels. By contrast, for the mammalian neonate the most important causes of its increased hypoxia tolerance may be just simple consequences of the comparatively undifferentiated state of the brain of the newborn, with its lower energy requirements, slower decline in ATP and lower excitability levels acting to delay depolarization.

Brain Na^+/K^+-ATPase activity in two anoxia tolerant vertebrates : Crucian carp and freshwater turtle

The crucian carp (Carassius carassius) and freshwater turtles (Trachemys scripta) are among the very few vertebrates that can survive extended periods of anoxia. The major problem for an anoxic brain is energy deficiency. In the brain, the Na+/K+-ATPase is the single most ATP consuming enzyme, being responsible for maintaining ion gradients. We here show that the Na+/K+-ATPase activity in the turtle brain is reduced by 31% in telencephalon and by 34% in cerebellum after 24 h of anoxia. Both changes were reversed upon reoxygenation. By contrast, the Na+/K+-ATPase activities were maintained in the anoxic crucian carp brain. These results support the notion that crucian carp and turtles use divergent strategies for anoxic survival. The fall in Na+/K+-ATPase activities displayed by the turtle is likely to be related to the strong depression of brain electric and metabolic activity utilized as an anoxic survival strategy by this species.

The Upregulation of Cognate and Inducible Heat Shock Proteins in the Anoxic Turtle Brain

Because heat shock proteins (HSPs) have an important protective function against ischemia/anoxia in mammalian brain, the authors investigated the expression of Hsp72 and Hsc73 in the anoxia-surviving turtle brain. Unlike the mammalian brain, high levels of Hsp72 were found in the normoxic turtle brain. Hsp72 levels were significantly increased by 4 hours of anoxia, remained constant until 8 hours, and then decreased to baseline at 12 hours. By contrast, Hsc73 was progressively increased throughout 12 hours of anoxia. This differential expression suggests different protective roles: Hsp72 in the initial downregulatory transition phase, and Hsc73 in maintaining neural network integrity during the long-term hypometabolic phase.

Adenosine, a “Retaliatory” Metabolite, Promotes Anoxia Tolerance in Turtle Brain

Contrary to what is found in most vertebrates, the brains of certain turtle species maintain ATP levels and ion homeostasis and survive prolonged anoxia. The hypothesis tested here is that the release of adenosine and its binding to A1 receptors are essential for this anoxic tolerance. Studies were conducted in the isolated turtle cerebellum, which did release adenosine to the extracellular space during anoxia. When adenosine receptor antagonists [theophylline, 8-cyclopentyltheophylline (CPT), or 8-cyclopentyl-1,3-dipropylxanthine (DPCPX)] were added to the superfusate under control conditions, they had no effect on extracellular potassium ion activity ([K+]o). During anoxia, however, these antagonists provoked maximal efflux of K+ (anoxic depolarization). Anoxic depolarization occurred earlier during anoxia with theophylline (a nonspecific adenosine receptor antagonist) than with CPT or DPCPX, which specifically block A1 receptors. Therefore, adenosine release and effects mediated by A1 receptors are essential to anoxia tolerance in turtle brain.

Voluntary diving metabolism and ventilation in the loggerhead sea turtle

Ventilation and gas exchange patterns were examined during voluntary dives in the loggerhead sea turtle Caretta caretta (L.) and contrasted with the changes that occur during forced submergence. Turtles spent 86% of their time submerged, with a mean dive length of 16.1 ± 6.0 (sd) min. During voluntary dives lasting 5–40 min, arterial blood PO2 fell to a minimum of 23 torr but turtles often surfaced before PO2 levels reached 50 torr. Arterial PCO2 increased typically < 10 torr while pH was held nearly constant, decreasing ⩽0.03-0.1U⪷dive−1. In voluntary submergence, aerobic limits were much longer than that of forced dives. The changes in blood acid-base status during voluntary submergence differed substantially from those reported in turtles forcibly submerged, and may account for the apparently reduced submersion endurance of sea turtles accidentally captured in shrimp trawls.