- Email: [email protected]
Effects of anaesthetics (halothane and MS-222) on crayfish, Astacus astacus
Aquaculture, 52 (1986) 2X3-217 Elsevier Science Publishers B.V., Amsterdam - Printed in The Netherlands
213
EFFECTS OF ANAESTHETICS (HALOTHANE AND MS-222) ON CRAYFISH, ASTACUS ASTACUS
J. OBRADOVIC Center for Marine Research
Zagreb, The Rudjer BolkoviE Institute, Zagreb (Yugoslavia)
(Accepted 21 December 1985)
ABSTRACT Obradovic, J., 1986. Effects of anaesthetics (halothane and MS-222) on crayfish, Astacus astacus. Aquaculture, 52: 213-217. The effects of an anaesthetic dispersive in air and usually applied in mammals (halothane, Hoechst) and of an anaesthetic soluble in water and applied in fish (MS-222, Sandoz) were investigated in experiments with crayfish, Astacus astacus (L.). Halothane was used at concentrations of 0.01, 0.06, 0.12, 0.5 and 1.0 vol. %, while MS-222 was applied in two concentrations, dissolved in the ratios of 1 : 1000 and 1 : 10 000. Halothane was most effective at 0.5 vol. %. The concentrations of MS-222 which were applied in our experiments had almost no effect on the crayfish.
INTRODUCTION
Anaesthetics have been in use for a long time in aquaculture because working with anaesthetized aquatic vertebrates was found to be advantageous in many technological operations, for example, fish spawning (Aganovic, 1979) and transportation of fish (Kindij, 1967). Numerous studies have been published about the effect of anaesthetics on fish (Laszlo, 1967; Bohl, 1968; Neimeth, 1968; Leitritz, 1969; Mann, 1969) but we have found no such reports on freshwater crayfish, although interest in the freshwater crayfish has increased in recent years not only because it is important in the biological cycle of inland waters (Brodskij, 1970; Momot and Jones, 1977; Cukeris and Terentjev, 1979; Obradovi& 1979) but even more so with regard to the possibility of producing crayfish for consumption (Romaire et al., 1977; Chien, 1980; ObradovG, 1983). A problem in crayfish breeding is how to examine thoroughly the roe which the female carries without losing a certain number of healthy roe as the result of the convulsive movements of the female during the examination. It is to be expected that with the development of astacoculture, the need for anaesthetics will also arise in other technological phases, and that anaesthetics will find a similar application in astacoculture as they now have in fish culture. It was therefore decided to investigate the effect of anaesthetics on crayfish. 0044-3486/86/$03.50
o 1986 Elsevier Science Publishers B.V.
214
MATERIAL
AND METHODS
Two anaesthetics (one generally used for aquatic organisms, the other for mammals) were chosen. The anaesthetics also differed in their mode of application (one applicable in aqueous solutions, the other dispersed in air). Halothane (Hoechst) was chosen as the anaesthetic dispersive in air and was applied to five different groups of crayfish in five different concentrations. Each group consisted of five test animals. The anaesthetic concentrations were prepared by soaking pieces of cotton wool in a measured quantity of halothane and subsequently covering them with a glass bell. We considered that the required concentrations of anaesthetic in the air under the glass bell (0.01, 0.06, 0.12, 0.5 and 1.0 vol. %) were obtained when the pieces of cotton wool were dry. A crayfish was placed under each bell. The period of exposure to the anaesthetic was determined separately for each group according to the behaviour of individual animals. MS-222 (Sandoz) was chosen as the water-soluble anaesthetic. It is the most frequently applied anaesthetic in aquaculture in Yugoslavia. MS-222 was applied in two concentrations in two experimental groups of crayfish, each group consisting of five animals. The experiment was conducted in glass baths containing a mixture of water and anaesthetic in the ratio of 1 : 10 000 and 1 : 1000. Both groups were exposed to the anaesthetic for a period of 30 min. In the experiment with halothane the crayfish were kept out of water for 30 min prior to exposure to the anaesthetic in order to achieve adaptation to the new conditions and to reduce the stress on the animals to the minimum, so as to allow clear interpretation of the reaction to the anaesthetic. In the experiment with MS-222 the crayfish were transferred directly from their water basin into the baths. The average length and weight of the crayfish in each group was 12.0 cm and 62 g respectively. The reactions of the crayfish were followed continuously, from the moment that anaesthesia began to take effect until the animals awoke and resumed their normal behaviour in the water and on land and reacted normally to stimulation. In view of the purpose of this experiment, special attention was paid to the reactions of the animals to stimulation and to the increased muscle tonus. RESULTS
The results of the experiment with halothane showed that (a) halothane was effective in crayfish, and (b) an effect on crayfish may be achieved by means of an anaesthetic dispersed in air. Fig. 1. showing reactions of the crayfish to various halothane concentrations indicates that even at a concentration of 0.01 vol. 7%the crayfish experienced a brief loss of consciousness, while at 0.06 vol. 7%this lasted from 9 to 13 min.
215
reactions Concentrations 0.01
--
0.06
---
0.12
-
time
in vol.%
in minutes
Fig. 1. Effects of various halothane concentrations on phases of narcosis in crayfish. Crayfish in precisely measured anaesthetic concentrations. 0 A Movements slowed down. Movements slowed down, vomiting. A, B Difficulties in the coordination of movements. Reaction to stimulation present, increased muscle tonus. B, Reaction to stimulation present, increased muscle tonus, vomiting. B, Reaction to stimulation absent, increased muscle tonus. B, c Comatose state. Anaesthetic eliminated. I -
The period necessary for the crayfish to recuperate and reach their normal state again depended on the concentration of the anaesthetic used. In these experiments this period ranged from 15 to 480 min. The phases which the exper~ent~ animals passed through in the course of ~aesthesia and afterwards also differed, depending on the doses received. Increased muscle tonus was observed with all concentrations of the anaesthetic except at 1.0 vol. %, difficulties in the coordination of movements were seen at 0.12 and 0.5 vol. %, vomiting at 0.12 vol. %, while the comatose state (state of complete numbness (coma) preceded and followed by lethargy) lasted for 3 h with a condensation of 1.0 vol. %. The results indicated that at 0.01 vol. % narcosis of short duration with a moderate reduction of reaction to stimuli was obtained and that the duration agreed with the increased muscle tonus, thus making the handling of the animals quite difficult. The state of unconsciousness lasted somewhat longer at a concentration of 0.06 vol. %, but was not considered satisfa~to~ because of the low intensity and the short duration. At 0.12 vol. % the duration of the impact of narcosis was satisfactory, but vomiting occurred. The best results were obtained at
216
0.5 vol. % because the period of increased muscle tonus during the narcosis was the shortest at this concentration. At the same time the narcosis lasts long enough to permit manipulation of the animals (Fig. 2.) Fig. 1. shows that the essential reactions to the anaesthetic in the animals investigated (such as loss of consciousness, reaction to stimuli, increased muscle tonus) were clearly different. This indicated that there was no linear increase in the nrtensity of these reactions in the crayfish due to higher halothane concentrations. The results of the experiment with MS-222 showed that it had no effect on the crayfish at a concentration of 1 : 10 000 and a duration of 30 min. However, at 1 : 1000 and a duration of 10 min a mild state of unconsciousness, lasting for approximately 10 min, developed. This suggested that the effects of anaesthesia tend to disappear during the course of the application. time in minutes
:t-
the time of q - anaesthesia
300 -
reaction, to
ELI - gk&llltlo”15 200 -
concentrations in vol.%
Fig. 2. The time of anaesthesia, influence and absence of reactions to stimulation.
DISCUSSION
A certain effect on the loss of consciousness, and possibly on the senses as well, was achieved by the application of the anaesthetic dispersed in air. However, application of the water-soluble anaesthetic had practically no effect on the crayfish. These results were contrary to our expectations since the physiology of respiration of the species in question is not too different from that of the aquatic vertebrates in which lower anaesthetic doses have given quite good results (Kindij, 1967). Thus the question remains open as to whether the unsuitability of the substance was due to the manner of application, as this would favour the assumption that it could not have penetrated into the organism by way of the gills (which seems acceptable
217
to us), or whether this substance simply does not affect the nervous system of the crayfish. The uncoordinated movements were probably caused by the increased tonus of certain muscles, but we do not know why they occurred only at higher concentrations of the anaesthetic. It is obvious that the anaesthetic provoked strong stimulation of the muscle tonus, manifested by contortion of all limbs and the tail. We may thus conclude that the reaction in the crayfish was more marked than the loss of consciousness at all halothane concentrations applied. With regard to the purpose of the experiments, we may conclude that application of an air-dispersive anaesthetic is more suitable for better manipulation of the crayfish. The best results were obtained in this case with halothane at a concentration of 0.5 vol. 76. ACKNOWLEDGEMENT
The author wishes to express her gratitude to the Self-Mangement Community of Interest for Scientific Research of SR Croatia for financial support.
REFERENCES AganoviE, M., 1979. Sahnonidne vrste riba i njihov uzgoj. Igkro “Svjetlost”, Sarajevo. Bohl, M., 1968. Uber die bet&bung von Laichforellen. Allg. Fisch. Ztg., 93: 124-125. Brodskij, S., 1970. Zaselit vodemi rakami. Ribovodstvo i Ribolovstvo, 4: 14. Chien, Y.R., 1980. Production of crayfish in rice fields. Prog. Fish-Cult., 42: 67-71. Cukeris, J.M. and Terentjev, A.L., 1979. Osobjenosti populaciji signaljnogo raka Pucifastacus Zeniusculus aklimatizirovanogo v izolirovanom vodoeme. Zoologi&eskij Zurnal, 58: 1733-1776. Kindij, Z., 1967. Transport Zive ribe, PriruEnik za slatkovodno ribarstvo. Agronomski glasnik, Zagreb. Laszlo, B., 1967. Halkabitasi kiserletek MS-222-Vel. Haleszad, 13 (60), 2 pp. Leitritz, E., 1969.Die Praxis der Forellenzucht. Paul Parey, Hamburg and Berlin. Mann, H., 1969. Uber den Einfluss eines Anaestheticum (MS-222 Sandoz) auf die Erbrutung von Forelleneiern. Fischwirt, 19 (8), 2 pp. Momot, W.T. and Jones, P.D., 1977. The relationship between biomass, growth rate and annual production in the crayfish, Orconectes virilis. Freshwater Crayfish, 3: 3-31. Neimeth, S., 1968. Halak kezelese es szailitasa boditott Qlapotban. Hal&ad, 13 (60), 3 PP. Obradovic, J., 1979. Problem smanjenja populacije rakova u slatkim vodama. Ribarstvo Jugoslavije, 1: 13. Obradovic, J., 1983. The possibilities of crayfish culture in Yugoslavia. Ribarstvo Jugoslavia, 38 : 56-57. Romaire, R.P., Forester, J.S. and Avault, J.W., Jr., 1977: Length-weight relationships of two commercially important crayfish of the genus Procambarus. Freshwater Crayfish, 3 : 463-470.
213
EFFECTS OF ANAESTHETICS (HALOTHANE AND MS-222) ON CRAYFISH, ASTACUS ASTACUS
J. OBRADOVIC Center for Marine Research
Zagreb, The Rudjer BolkoviE Institute, Zagreb (Yugoslavia)
(Accepted 21 December 1985)
ABSTRACT Obradovic, J., 1986. Effects of anaesthetics (halothane and MS-222) on crayfish, Astacus astacus. Aquaculture, 52: 213-217. The effects of an anaesthetic dispersive in air and usually applied in mammals (halothane, Hoechst) and of an anaesthetic soluble in water and applied in fish (MS-222, Sandoz) were investigated in experiments with crayfish, Astacus astacus (L.). Halothane was used at concentrations of 0.01, 0.06, 0.12, 0.5 and 1.0 vol. %, while MS-222 was applied in two concentrations, dissolved in the ratios of 1 : 1000 and 1 : 10 000. Halothane was most effective at 0.5 vol. %. The concentrations of MS-222 which were applied in our experiments had almost no effect on the crayfish.
INTRODUCTION
Anaesthetics have been in use for a long time in aquaculture because working with anaesthetized aquatic vertebrates was found to be advantageous in many technological operations, for example, fish spawning (Aganovic, 1979) and transportation of fish (Kindij, 1967). Numerous studies have been published about the effect of anaesthetics on fish (Laszlo, 1967; Bohl, 1968; Neimeth, 1968; Leitritz, 1969; Mann, 1969) but we have found no such reports on freshwater crayfish, although interest in the freshwater crayfish has increased in recent years not only because it is important in the biological cycle of inland waters (Brodskij, 1970; Momot and Jones, 1977; Cukeris and Terentjev, 1979; Obradovi& 1979) but even more so with regard to the possibility of producing crayfish for consumption (Romaire et al., 1977; Chien, 1980; ObradovG, 1983). A problem in crayfish breeding is how to examine thoroughly the roe which the female carries without losing a certain number of healthy roe as the result of the convulsive movements of the female during the examination. It is to be expected that with the development of astacoculture, the need for anaesthetics will also arise in other technological phases, and that anaesthetics will find a similar application in astacoculture as they now have in fish culture. It was therefore decided to investigate the effect of anaesthetics on crayfish. 0044-3486/86/$03.50
o 1986 Elsevier Science Publishers B.V.
214
MATERIAL
AND METHODS
Two anaesthetics (one generally used for aquatic organisms, the other for mammals) were chosen. The anaesthetics also differed in their mode of application (one applicable in aqueous solutions, the other dispersed in air). Halothane (Hoechst) was chosen as the anaesthetic dispersive in air and was applied to five different groups of crayfish in five different concentrations. Each group consisted of five test animals. The anaesthetic concentrations were prepared by soaking pieces of cotton wool in a measured quantity of halothane and subsequently covering them with a glass bell. We considered that the required concentrations of anaesthetic in the air under the glass bell (0.01, 0.06, 0.12, 0.5 and 1.0 vol. %) were obtained when the pieces of cotton wool were dry. A crayfish was placed under each bell. The period of exposure to the anaesthetic was determined separately for each group according to the behaviour of individual animals. MS-222 (Sandoz) was chosen as the water-soluble anaesthetic. It is the most frequently applied anaesthetic in aquaculture in Yugoslavia. MS-222 was applied in two concentrations in two experimental groups of crayfish, each group consisting of five animals. The experiment was conducted in glass baths containing a mixture of water and anaesthetic in the ratio of 1 : 10 000 and 1 : 1000. Both groups were exposed to the anaesthetic for a period of 30 min. In the experiment with halothane the crayfish were kept out of water for 30 min prior to exposure to the anaesthetic in order to achieve adaptation to the new conditions and to reduce the stress on the animals to the minimum, so as to allow clear interpretation of the reaction to the anaesthetic. In the experiment with MS-222 the crayfish were transferred directly from their water basin into the baths. The average length and weight of the crayfish in each group was 12.0 cm and 62 g respectively. The reactions of the crayfish were followed continuously, from the moment that anaesthesia began to take effect until the animals awoke and resumed their normal behaviour in the water and on land and reacted normally to stimulation. In view of the purpose of this experiment, special attention was paid to the reactions of the animals to stimulation and to the increased muscle tonus. RESULTS
The results of the experiment with halothane showed that (a) halothane was effective in crayfish, and (b) an effect on crayfish may be achieved by means of an anaesthetic dispersed in air. Fig. 1. showing reactions of the crayfish to various halothane concentrations indicates that even at a concentration of 0.01 vol. 7%the crayfish experienced a brief loss of consciousness, while at 0.06 vol. 7%this lasted from 9 to 13 min.
215
reactions Concentrations 0.01
--
0.06
---
0.12
-
time
in vol.%
in minutes
Fig. 1. Effects of various halothane concentrations on phases of narcosis in crayfish. Crayfish in precisely measured anaesthetic concentrations. 0 A Movements slowed down. Movements slowed down, vomiting. A, B Difficulties in the coordination of movements. Reaction to stimulation present, increased muscle tonus. B, Reaction to stimulation present, increased muscle tonus, vomiting. B, Reaction to stimulation absent, increased muscle tonus. B, c Comatose state. Anaesthetic eliminated. I -
The period necessary for the crayfish to recuperate and reach their normal state again depended on the concentration of the anaesthetic used. In these experiments this period ranged from 15 to 480 min. The phases which the exper~ent~ animals passed through in the course of ~aesthesia and afterwards also differed, depending on the doses received. Increased muscle tonus was observed with all concentrations of the anaesthetic except at 1.0 vol. %, difficulties in the coordination of movements were seen at 0.12 and 0.5 vol. %, vomiting at 0.12 vol. %, while the comatose state (state of complete numbness (coma) preceded and followed by lethargy) lasted for 3 h with a condensation of 1.0 vol. %. The results indicated that at 0.01 vol. % narcosis of short duration with a moderate reduction of reaction to stimuli was obtained and that the duration agreed with the increased muscle tonus, thus making the handling of the animals quite difficult. The state of unconsciousness lasted somewhat longer at a concentration of 0.06 vol. %, but was not considered satisfa~to~ because of the low intensity and the short duration. At 0.12 vol. % the duration of the impact of narcosis was satisfactory, but vomiting occurred. The best results were obtained at
216
0.5 vol. % because the period of increased muscle tonus during the narcosis was the shortest at this concentration. At the same time the narcosis lasts long enough to permit manipulation of the animals (Fig. 2.) Fig. 1. shows that the essential reactions to the anaesthetic in the animals investigated (such as loss of consciousness, reaction to stimuli, increased muscle tonus) were clearly different. This indicated that there was no linear increase in the nrtensity of these reactions in the crayfish due to higher halothane concentrations. The results of the experiment with MS-222 showed that it had no effect on the crayfish at a concentration of 1 : 10 000 and a duration of 30 min. However, at 1 : 1000 and a duration of 10 min a mild state of unconsciousness, lasting for approximately 10 min, developed. This suggested that the effects of anaesthesia tend to disappear during the course of the application. time in minutes
:t-
the time of q - anaesthesia
300 -
reaction, to
ELI - gk&llltlo”15 200 -
concentrations in vol.%
Fig. 2. The time of anaesthesia, influence and absence of reactions to stimulation.
DISCUSSION
A certain effect on the loss of consciousness, and possibly on the senses as well, was achieved by the application of the anaesthetic dispersed in air. However, application of the water-soluble anaesthetic had practically no effect on the crayfish. These results were contrary to our expectations since the physiology of respiration of the species in question is not too different from that of the aquatic vertebrates in which lower anaesthetic doses have given quite good results (Kindij, 1967). Thus the question remains open as to whether the unsuitability of the substance was due to the manner of application, as this would favour the assumption that it could not have penetrated into the organism by way of the gills (which seems acceptable
217
to us), or whether this substance simply does not affect the nervous system of the crayfish. The uncoordinated movements were probably caused by the increased tonus of certain muscles, but we do not know why they occurred only at higher concentrations of the anaesthetic. It is obvious that the anaesthetic provoked strong stimulation of the muscle tonus, manifested by contortion of all limbs and the tail. We may thus conclude that the reaction in the crayfish was more marked than the loss of consciousness at all halothane concentrations applied. With regard to the purpose of the experiments, we may conclude that application of an air-dispersive anaesthetic is more suitable for better manipulation of the crayfish. The best results were obtained in this case with halothane at a concentration of 0.5 vol. 76. ACKNOWLEDGEMENT
The author wishes to express her gratitude to the Self-Mangement Community of Interest for Scientific Research of SR Croatia for financial support.
REFERENCES AganoviE, M., 1979. Sahnonidne vrste riba i njihov uzgoj. Igkro “Svjetlost”, Sarajevo. Bohl, M., 1968. Uber die bet&bung von Laichforellen. Allg. Fisch. Ztg., 93: 124-125. Brodskij, S., 1970. Zaselit vodemi rakami. Ribovodstvo i Ribolovstvo, 4: 14. Chien, Y.R., 1980. Production of crayfish in rice fields. Prog. Fish-Cult., 42: 67-71. Cukeris, J.M. and Terentjev, A.L., 1979. Osobjenosti populaciji signaljnogo raka Pucifastacus Zeniusculus aklimatizirovanogo v izolirovanom vodoeme. Zoologi&eskij Zurnal, 58: 1733-1776. Kindij, Z., 1967. Transport Zive ribe, PriruEnik za slatkovodno ribarstvo. Agronomski glasnik, Zagreb. Laszlo, B., 1967. Halkabitasi kiserletek MS-222-Vel. Haleszad, 13 (60), 2 pp. Leitritz, E., 1969.Die Praxis der Forellenzucht. Paul Parey, Hamburg and Berlin. Mann, H., 1969. Uber den Einfluss eines Anaestheticum (MS-222 Sandoz) auf die Erbrutung von Forelleneiern. Fischwirt, 19 (8), 2 pp. Momot, W.T. and Jones, P.D., 1977. The relationship between biomass, growth rate and annual production in the crayfish, Orconectes virilis. Freshwater Crayfish, 3: 3-31. Neimeth, S., 1968. Halak kezelese es szailitasa boditott Qlapotban. Hal&ad, 13 (60), 3 PP. Obradovic, J., 1979. Problem smanjenja populacije rakova u slatkim vodama. Ribarstvo Jugoslavije, 1: 13. Obradovic, J., 1983. The possibilities of crayfish culture in Yugoslavia. Ribarstvo Jugoslavia, 38 : 56-57. Romaire, R.P., Forester, J.S. and Avault, J.W., Jr., 1977: Length-weight relationships of two commercially important crayfish of the genus Procambarus. Freshwater Crayfish, 3 : 463-470.