Summary

Twenty one spiny dogfish were captured, handled, and transported a short distance. Sixteen were treated with azaperone (4mg/kg) and showed a significant Increase in blood glucose levels when compared to 5 similarly treated with seawater. Tranquilized animals showed little resistance to recapture or routine sampling procedures. All animals treated with azaperone exhibited normal swimming behavior. It is postulated that azaperone acts directly to release glucose to the blood and by blocking effects of stress-induced catecholamines, and indirectly by blocking dopaminergic receptors of the ascending reticular activating system and mesolimbic pathways. It is suggested that this agent be used In capture and transport of larger pelagic species.

Introduction

Capture, transport, and confinement of sharks for public display is often unsuccessful. Many species do not easily adapt to captivity or manual restraint. Sharks often succumb to stress imposed by routine handling required for transport or blood sample procurement. Maintenance of sharks in captivity, especially of the larger, pelagic species, meets with failure even though public interest is keen regarding the most delicate species.

A satisfactory regimen for tranquilization of sharks has been sought. Many chemical agents have been used with varying results, including: MS-222, a tranquilizer used in teleost fish and amphibians;¹ quinaldine, an agent used for immobilization of marine invertebrates and vertebrates;¹³ Saffan,^R a steroidal anesthetic commonly used in felines;⁷ and halothane.⁵ Physical methods of restraint have included attempts at tonic immobilization, confinement in portable transport containers, cold shock, and carbon dioxide poisoning.

The above methods have proved unsuccessful for the most part. The purpose of this study was to find a means of chemical restraint which would be easy to administer yet not initiate or exacerbate the stress of capture and routine handling. Common drugs used in veterinary medicine for immobilization and sedation are often contraindicated in animals showing cardiovascular embarrassment or collapse. A regimen suggested by Stoskopf¹⁵ using ketamine and xylazine is suitable for animals adjusted to captivity. However, it is widely assumed that newly-caught sharks exhibit signs of "shock", a state in which xylazine is contraindicated.

Sharks require some muscular contractions to assist venous return to the heart. Anxiolytic drugs which allow normal motor activity and respiration are highly desirable in captive sharks. In addition, the ideal agent should assist adaptation to the confined environment. Suppression of escape behavior and self-induced trauma following release would be major advantages. Azaperone (Stresnil Pitman-Moore) was selected for this study. Azaperone is a butyrophenone tranquilizer. It is used in the swine industry to control aggressiveness in pigs when litters ate intermixed. A single dose is administered IM, and fighting between the pigs is decreased. Use of this drug has shown no contraindications. Azaperone "tames" aggressive animals by block Ing dopaminergic receptors of the mesolimbic and nigrostriatal pathways and by decreasing the activity of the ascending reticular activating system (ARAS). This drug thusly reduces response to the environment without motor Impairment or sedation.

Spiny dogfish (Squalus acanthia) were selected as experimental subjects. They are readily available in the Tacoma area, are easily maintained in display, yet show a variety of capture and captivity-related syndromes which may be evaluated easily. Preliminary studies showed the most efficacious application of azaperone to be directly over the gill rather than by injection. A tuberculin syringe was introduced through a gill slit and the drug deposited on the gill filaments.

Methods

Ten male and eleven female spiny dogfish were used in this study. Weights of the animals ranged from 1-5 kg. All had been captured 7 to 10 days prior to the study. The animals were housed in the main display tank at the Point Defiance Zoo and Aquarium, Tacoma, WA. All subjects were removed from the water with a dip net. Each was manually restrained and a 12 ml blood sample taken from the dorsal caudal vein with a syringe and 18 ga. needle. A tuberculin syringe was used to place 4 mg/kg azaperone through the gill slit and directly on the gills of 16 animals. Five received only seawater as sham treatment. The slits were held closed for several seconds. The pectoral fins were punched for identification and the animal was placed in a container for transport to an outdoor tank. No animal was out of water for more than 30 seconds nor restrained and transported for more than 3 minutes. The sharks were not disturbed for 4 hours following treatment. Periodic observations were made of each animal noting behavioral changes and activity.

Four hours after azaperone administration, each dogfish was netted and a 5 ml blood sample was taken. The following blood values were obtained from control samples: glucose, BUN, creatinine kinase, cholesterol, total protein, albumin, calcium phosphorus, alkaline phosphatase, SGPT, sodium, potassium, and chloride. Only glucose levels were determined from trial samples. Data was evaluated with the student's t-test with P < 0.05 considered significant.

Results

One animal died during capture and sampling. The remaining twenty animals were alive, feeding, and healthy 3 months following the trial. All 21 subjects resisted initial capture and blood sampling strenuously. Those treated with azaperone appeared calm upon release to the outdoor pool. The 5 animals treated with seawater showed excited swimming behavior, thrashing and planing across the surface in a typical agitated manner. They rammed the sides and feeding deck often. Treated animals swam normally and did not show escape behavior.

Recapture of tranquilized animals was easily accomplished, and blood samples taken with little or no restraint. Nontranquilized animals were difficult to capture, restrain and venipuncture was hampered.

Treated animals readily accepted food the day following treatment. Untreated animals did not feed for several days.

Animals treated with azaperone are shown separately from those treated with seawater. The control values of glucose levels between the two groups were riot significantly different with control glucose being 28±14 mg/dl for both groups. Trial glucose levels were significantly different at the p=.01 level. Trial value- was 41±11 mg/dl for animals which received azaperone, and 24±5 mg/dl for those receiving seawater. There was no difference between trial and control glucose levels in the group treated with seawater.

Discussion

The response to catecholamines in the spiny dogfish has been measured using glucose levels as indirect evaluation.⁴ Dogfish show a response similar to that seen in mammals: a steep rise in glucose values which peaks at 3-4 hours, and a gradual decline over the next 24 hours. The mechanism of glycolysis has not been established. Very little glycogen (approximately 1000 cal worth) is present in the liver of spiny dogfish, as the majority of hepatic function is related to fat metabolism and maintenance of neutral buoyancy.¹⁰ It has been postulated that glucose may arise from anabolic or catabolic pathways utilizing tissue proteins.⁴ Liver glycogen has been shown to be unresponsive to ACTH or corticosteroids, which respectively elevated glucose levels. Administration of 4 mg/kg azaperone across the gills of spiny dogfish causes a rise in glucose levels.^3,4,12 Several modes of action may be postulated. The drug may be acting directly to cause release of glucose. No significant glucocorticoid activity has been shown from the secretions of the internal (adrenal) organ.¹⁰ Azaperone may be functioning as such.

Escape behavior identical to that in spiny dogfish has been noted in the nurse shark (ginglymostoma cirratum). This behavior is elicited by electrical stimulation of specific regions of the brain. Demski² suggests that this area may be equivalent to the tetrapod limbic system. Suppression of escape behavior with azaperone is consistent with its action of mesolimbic dopaminergic blockade.

The drug may also be acting directly on the ARAS, decreasing awareness of stress in the individual and thereby decreasing metabolism of glucose previously released into the blood. Nontreated animals may be metabolizing glucose released by stressful activity, and show an apparent decrease in glucose from control levels. Or, they may have been so exhausted from capture and confinement that glucose reserves were depleted and the fish were unable to respond.

Azaperone has no harmful side effects. All animals treated with the drug returned to normal behavior within 24 hours. They fed readily; unusual in dogfish, which often starve to death in captivity over several months. Some decrease in escape behavior, over several weeks, was noted by the aquarists, who claimed fewer of the treated animals indulged in self traumatizing activity.

Conclusions

The ideal drug for the capture and confinement of sharks would decrease anxiety, yet allow normal swimming, aeration of the gills, and cardiovascular function. Azaperone at 4 mg/kg applied to the gills apparently fulfills these requirements in the spiny dogfish.

These effects of azaperone may be especially important in the warm-water species which are more prone to panic, aggression, and self-induced trauma. Specimens of the great white shark (Carchardon carcharius) have been successfully transported, nursed through capture-related shock, but have died upon release in display tanks.^8,11 Valuable specimens such as these may benefit from repeated application of azaperone until they adjust to the new environment enabling many aquaria to obtain and maintain these species that are of intense public interest.

Acknowledgements

I wish to thank many people who have assisted me and given advice: John Rupp, Dr. Leroy Gallagher, Dr. Tom Riebold, Dr. Michael Stoskopf, the entire staff of the Veterinary Diagnostic Labs at WSU and OSU, and the staff at the Point Defiance Zoo and Aquarium.

References

1.  Campbel, G.D. and Davies, D.H.: The effect of MS-222 on the elasmobranch electrocardiograph. Nature 198:302, 1963.

2.  Demski, E.M.: Electrical stimulation of the shark brain. Amer. Zool. 17:487-500, 1977.

3.  deRoos, R. and deRoos, C.C.: Comparative effects of the pituitary adrenal axis and catecholamines on carbohydrate metabolism in elasmobranch fish. Gen. Comp. Endocrinology Suppl. 3:192-197, 1972.

4.  Elevation of plasma glucose levels by catecholamines in elasmobranch fish. Gen. Comp. Endocrinol. 34:447-452, 1978.

5.  Dunn, R.F. and Koester, D.M.: A short review of the use of anesthetics in elasmobranchs with particular interest on the use of halothane oxygen-nitrous oxide. Recent Advances In Captive Biology of Sharks, Johns Hopkins Press, Baltimore, MD., 1985.

6.  Gruber, S.H. and Keyes, R.S.: Keeping sharks for research, Aquarium Systems, A.D. Hawkins, ed., Academic Press, N.Y., 1981, pp.373-402.

7.  Harvey, B.: New sedation techniques for sharks: Saffan administered by an underwater dart gun. Ann. Proc. American Elasmobranch Soc., U. of Victoria Press, Victoria, B.C., 1986.

8.  Hewitt, J.: Personal communication. Steinhart Aquarium, Golden Gate Park, San Francisco, 1986.

9.  Jones, B.C.: Feeding behavior and survival of Pacific spiny dogfish in captivity. Progr. Fish Culturist 40(4):157, 1978.

10. Martini, F.H.: The effects of fasting confinement on Squalus acanthias Sensory Biology of Sharks, Skates, and Rays. GPO US Navy res. pp.608-646, 1978.

11. Mckosker, J.: Personal communication, Steinhart Aquarium, Golden Gate Park, San Francisco. 1986.

12. Patent, G.J.: Comparison of some hormonal effects on carbohydrate metabolism in an elasmobranch acanthias and a holocephalan (Hydrolagus collei). Gen. Comp. Endocrinol. 16:535-554, 1970.

13. Rupp, J.: Personal communication. Pt. Defiance Aquarium, Tacoma, WA. 1986.

14. Stoskopg, M.K.: Personal communication. National Aquarium, Baltimore, MD. 1985.

15. Preliminary notes and anesthesia of captive sharks. (in press) 1986.