Physiology & Behavior, Vol. 22, pp. 25-29. PergamonPress and Brain ResearchPubl., 1979. Printed in the U.S.A.
Can Halothane Anaesthesia Have Any Aversive Effects on the Rat? GERARD SCHMALTZ Laboratoire de Psychophysiologie, Universit$ de Lille 1, B P 36, 59650 Villeneuve d ' A s c q , France
( R e c e i v e d 3 M a r c h 1978) SCHMALTZ, G. Can halothane anaesthesia have any aversive effects on the rat? PHYSIOL. BEHAV. 2,2(1) 25-29, 1979.--Rats (3x2 groups) were put in a Y maze and submitted to one of the f o ~ 3 Wetresm~ts OPT): 0, 3, I0 uncontrollable presboeks (PS). Each PT, administered to 2 groups, was immediately foUowed either by Imiothlmemutesthetiz/ng (groups M,,_,, M:,_., M,,_,0 or sham anaesthetizing (groups M,, M.~,M,o). One day later, the 6 Woups underwent active avoidance conditioning in the Y maze. The overall escape performanee Ofgroup M~did not differ from that Ofjroup M,,. By contrast, a marked impairment was observed in the M,,, group, which demonstrates both the effectiveness and the good memorisation of the most aversive PT (10 PS). The acquisition of group M,,_,, was as much Lqq3airedas that oflgoup M.,. Consequently, we can conclude that halothane has no amnesic effects. The performances of groups Mo-, and M~_, were inferior to those of groups M,, and M:,, respectively. These last results are discussed in terms of aversive effects of halothane. Rat
Preshocks and haiothane anaesthesia
Aversive effects
Avoidance conditioning
SCHMALTZ, G. Une anesthisie au fluothane a-t-elle des effets aversifs chez le rat? PHYSIOL. BEHAV. 22(1) 25-29, 1979.--Des rats (3x2 groupes) ref~vem, dans un labyrimhe en Y, I'un des 3 pr~traitements (FI') suivants: 0, 3, 10 checs ¢gectriques incontrblables (CI). Chaque IT, administr~ h 2 groupes de sujets, est imm~iatement suivi, soit par une anesth~sie au fluothane (groupes t}A, 3-A, It}A), soit par une anesth~sie simukbe(groupes O-AS, 3-AS, It}AS). Unjour plus tard, les 6 gronpes effectuent un conditionnement d'~vitement actif dans le labyrinthe en Y. La performance O~.hsppement) du Woupe 3--AS ne di/~h~ pes de celle du groupe t}AS. Par contre, u u nette perlturbatkm s'observe ~ le groupe 10.-AS, ce qoi d6montre it la fois l'efl]caci~ et ht bonne n~morimt/on du PT le plus aversif (I0 CI). L'aPlmmlistqle du groupe It}A est aussi perturb6 clue ceiui du groupe 10AS. Nous pouvons en conclure qne le fluothane n'a pes d'effets amn6siques. Les performances des groupes 0--A et 3-A sont, respectivement, inf6rieures h ceIles des groupes 0-AS et 3-AS. Ces derniers r~sultats sont interpr~t~s en termes d'effets aversifs du fluothane. Rat
Chocs 61ectriques et anesth~sie au fluothane
Effets aversifs
H A L O T H A N E seems to produce fewer adverse reactions in the cardio-vascular and respiratory systems than other anaesthetics [8]. It is administered by inhalation and acts very rapidly but also wears off rapidly. In Wistar strain rats, for example, narcosis occurs in less than a minute, minimizing strusgling, and yet after 2 hr under anaesthetic, the animaps behaviour returns to normal within a few minutes [2]. The administration of this treatment thus appears not to be traumatic for the animals. Partly because of these advantages, halothane has been used to interrupt the consolidation phase of memory formation [2, 4, 6, 7]. The essential evidence which permits one to draw this conclusion is the presence of a temporal gradient of amnesia, that is, the shorter the time interval between learning and treatment, the greater the impairment [2, 4, 13]. For details of the experimental results obtained on rats see the excellent reviews by Bloch [3] and Deweer [5], both concerned with the consolidation of memory. In addition to the anmesic effects of halothane, and despite its apparent inoffensiveness, it is nevertheless possible that this anaesthetic also has aversive properties. This hy-
Conditionnement d'C~vitement
pothesis has already received some confirmation, and the work of Verrier-Oisquet et al. [11], Alexinsky and Chaponthier [1] on rats and Zirgkin et al. [13] on chicks has indicted that iwlo~ane may have not only ~ but also aversive effects. The object of the present experiment is to investigate the possible aversive effects of an ludotlmae anaesthesia on the rat and not to distinguish between aversive and amnesic effects. We start from the result of a previous study [9] which has shown that aversive uncontrollable shocks administered to rats as a pretreatment (preshocks) may interfere with the later acquisition of an avoidance task. In the present study, all the rats first received preshocks in a Y maze. Then, half of the animals was immediately anaesthetized with halothane. The possible effects of halothane will be more easily detectable when there is no delay between preshocks and anaesthesht. One day after these pretreatments, all the animals were given an avoidance learning session in the Y maze. If the anaesthesia has aversive properties, then it should increase the impairment produced by the preshocks alone so that the performance of the preshocked-anaesthetized rats should be inferior to that of
C o p y r i g h t '~ 1979 Brain R e s e a r c h Publications Inc.~0031-9384/79/010025-05502.00/0
26
SCHMALTZ
the preshocked ones. In short, one should be able to detect the aversive effects of halothane anaesthesia, if any, by examining the performance of the animals during the avoidance conditioning session. METHOD Animals The animals were 85 male Sprague-Dawley rats weighing between 285-315 g at the start of the experiment. All the animals were housed individually, with ad lib food and water available in their home cages. They were handled once each morning during the week before the beginning of the experiment. Apparatus The apparatus used was a Y-nmze with an adjoining start box. All the walls were wooden. The 3 arms of the maze formed 120" angles, and each arm was 43 cm long, 12 cm wide and 30 cm high. The centrld alley was soperated from the start box by a ~ door, while each of the lateral arms had a slidlnll door 30 cm away from the ~ end. These doors ~ the e ~ t e r to isolate ~ animal in the goal box. The floor of the wmze, including the start box, consisted of parallel brass rods 3 nun in dian~ter lying at 10 m m intervals. This grid was connected to a constant current stimulator. The entire apparatus was covered with Perspex lids. Procedure The 85 animals were divided at random into 8 groups (N= 10, except for one Stoup with N=IS). For each animal, the experiment involved 2 sessions: pretreatment, then avoidance conditioning. Session 1: Pretreatment Non-anaesthetized groups. In 3 of the groups (M., M:j and M,o) the animals were placed individually in the maze where they remained for 30 win. They could not enter the start box, as door was kept closed. During this half hour the rats in these groups received, respectively, 0, 3 or 10 aversive preshocks (1,5 mA in pulsed train~--0.1 sec on; 0,4 sac oft). These l ~ k s (PS) were inesc,e p ~ and u n ~ e d , the onset, duration and intensity being controlled by the experimenter. The fact that the PS were uncontrollable considerably increased the aversiveness of the situation [10]. Immediately after the 30 rain spent in the maze, the aninmls were given sham anaesthesia (see below) then returned to their home cages. This sham a n a e s t h u was also given to rats in a control group Co, which were never placed in the maze, but simply renmined in their home cages. Anaesthetized groups. Groups C._j,, M._., M:,_,, and M,o-. received exactly the same pretreamumts (PS) as their counterparts described above, except that a FFdotex Mark 2 delivered a mixture of equal parts o f e l r and oxygen (flow 2 l/rain) with a 1,5% concentration of halothaae. The mixture passed through a Ptexigias box (25 cm×25 cm×25 cm), escaping through 4 small h ~ e s in the lid. Each rat was placed individually in this box for 60 rain. The average time to achieve narcosis was 40 sex. After spending an hour under the influence of anaesthetic, the unconscious animal was removed and placed in its home cage where it recovered fully
within 10 to 15 min. After waking there was no obvious sign of impairment in gross behaviour. The sham anaesthesia was performed in the same box, but there was no halothane in the air-oxygen mixture, and the time spent in the box was only 40 sec. Session 2: Avoidance Conditioning The active avoidance conditioning involved 15 trials in the Y-maze and took place one day after the first session. The same procedure, as previously described [9], was used for all the rats. The rat was placed in the start box at the beginning of the session. After 1 rain, the door was opened, allowing access to the maze alleys. Five sec later, the rat received footshock with the same characteristics as the pretreatment shock, except that now it could be controlled by the animal. The shock stopped when the animal, after reaching the end of one arm (first choice), turned round and went to the end of the other arm beyond the sliding door (second choice), This sliding door was then closed and the animal was left in the goal box for 30 sex before being placed in the start box for the I rain intertrial period. During the first trial it was thus necessary for the rat to cover the entire maze before escaping the aversive stimulus (escape response). During the following trials, the goal box (second choice on the first trial) was no longer electrified. The rat could now either make an escape response by entering the goal box after the shock had begun, or make a conditioned avoidance response (CAR) by entering the goal box directly--i.e., without making any spatial discrimination error--before the onset of the shock. Some animals, especially in the early stages of learning, did not escape the shock. In this case the experimenter intervened. During the first trial if, after a 20 sex stimulation period, the rat was still in the start box, it was pushed into the central alley and door was closed. If after 90 sex the animal had not made the escape response it was pushed into a goal box for 10 sex. Therefore shock could not last longer than I00 sex on any trial. The location of the goal box depended on the previous behaviour of the animal. The goal box was either the unexplored arm, or an arm was chosen at random if the rat had not explored either arm. During the remaining trials, the procedure was the same except that the goal box was already determined. RESULTS Behavioural Observations During Pretreatment While the rats were given these 3 uncontrollable PS, their most frequent response was flight. In some a n i ~ (8/20), freezing appeared on the third PS. Among those animals which were submitted to 10 uncontrollable PS, the same pattern was observed during the early shocks but freezing then increase in frequency. After the fifth PS, the animals froze consistently in response to the aversive stimulus. Avoidance Conditioning in Non-anaesthetized Groups During the active avoidance conditioning we recorded the duration of stimulation on each trial (range: 0-100 sec)for each rat and also the total duration of stimulation summed over all 15 trials. This enabled us to calculate, respectively, the mean duration of stimulation (MDS) per trial for a given group and the mean total duration of stimulation (MTDS) for the group. A Preshock×Anaesthesia analysis of variance
AVERSIVE EFFECTS OF HALOTHANE
27
demonstrated a statistically significant preshock effect, F(2,59)= 14.47, p
Can Halothane Anaesthesia Have Any Aversive Effects on the Rat? GERARD SCHMALTZ Laboratoire de Psychophysiologie, Universit$ de Lille 1, B P 36, 59650 Villeneuve d ' A s c q , France
( R e c e i v e d 3 M a r c h 1978) SCHMALTZ, G. Can halothane anaesthesia have any aversive effects on the rat? PHYSIOL. BEHAV. 2,2(1) 25-29, 1979.--Rats (3x2 groups) were put in a Y maze and submitted to one of the f o ~ 3 Wetresm~ts OPT): 0, 3, I0 uncontrollable presboeks (PS). Each PT, administered to 2 groups, was immediately foUowed either by Imiothlmemutesthetiz/ng (groups M,,_,, M:,_., M,,_,0 or sham anaesthetizing (groups M,, M.~,M,o). One day later, the 6 Woups underwent active avoidance conditioning in the Y maze. The overall escape performanee Ofgroup M~did not differ from that Ofjroup M,,. By contrast, a marked impairment was observed in the M,,, group, which demonstrates both the effectiveness and the good memorisation of the most aversive PT (10 PS). The acquisition of group M,,_,, was as much Lqq3airedas that oflgoup M.,. Consequently, we can conclude that halothane has no amnesic effects. The performances of groups Mo-, and M~_, were inferior to those of groups M,, and M:,, respectively. These last results are discussed in terms of aversive effects of halothane. Rat
Preshocks and haiothane anaesthesia
Aversive effects
Avoidance conditioning
SCHMALTZ, G. Une anesthisie au fluothane a-t-elle des effets aversifs chez le rat? PHYSIOL. BEHAV. 22(1) 25-29, 1979.--Des rats (3x2 groupes) ref~vem, dans un labyrimhe en Y, I'un des 3 pr~traitements (FI') suivants: 0, 3, 10 checs ¢gectriques incontrblables (CI). Chaque IT, administr~ h 2 groupes de sujets, est imm~iatement suivi, soit par une anesth~sie au fluothane (groupes t}A, 3-A, It}A), soit par une anesth~sie simukbe(groupes O-AS, 3-AS, It}AS). Unjour plus tard, les 6 gronpes effectuent un conditionnement d'~vitement actif dans le labyrinthe en Y. La performance O~.hsppement) du Woupe 3--AS ne di/~h~ pes de celle du groupe t}AS. Par contre, u u nette perlturbatkm s'observe ~ le groupe 10.-AS, ce qoi d6montre it la fois l'efl]caci~ et ht bonne n~morimt/on du PT le plus aversif (I0 CI). L'aPlmmlistqle du groupe It}A est aussi perturb6 clue ceiui du groupe 10AS. Nous pouvons en conclure qne le fluothane n'a pes d'effets amn6siques. Les performances des groupes 0--A et 3-A sont, respectivement, inf6rieures h ceIles des groupes 0-AS et 3-AS. Ces derniers r~sultats sont interpr~t~s en termes d'effets aversifs du fluothane. Rat
Chocs 61ectriques et anesth~sie au fluothane
Effets aversifs
H A L O T H A N E seems to produce fewer adverse reactions in the cardio-vascular and respiratory systems than other anaesthetics [8]. It is administered by inhalation and acts very rapidly but also wears off rapidly. In Wistar strain rats, for example, narcosis occurs in less than a minute, minimizing strusgling, and yet after 2 hr under anaesthetic, the animaps behaviour returns to normal within a few minutes [2]. The administration of this treatment thus appears not to be traumatic for the animals. Partly because of these advantages, halothane has been used to interrupt the consolidation phase of memory formation [2, 4, 6, 7]. The essential evidence which permits one to draw this conclusion is the presence of a temporal gradient of amnesia, that is, the shorter the time interval between learning and treatment, the greater the impairment [2, 4, 13]. For details of the experimental results obtained on rats see the excellent reviews by Bloch [3] and Deweer [5], both concerned with the consolidation of memory. In addition to the anmesic effects of halothane, and despite its apparent inoffensiveness, it is nevertheless possible that this anaesthetic also has aversive properties. This hy-
Conditionnement d'C~vitement
pothesis has already received some confirmation, and the work of Verrier-Oisquet et al. [11], Alexinsky and Chaponthier [1] on rats and Zirgkin et al. [13] on chicks has indicted that iwlo~ane may have not only ~ but also aversive effects. The object of the present experiment is to investigate the possible aversive effects of an ludotlmae anaesthesia on the rat and not to distinguish between aversive and amnesic effects. We start from the result of a previous study [9] which has shown that aversive uncontrollable shocks administered to rats as a pretreatment (preshocks) may interfere with the later acquisition of an avoidance task. In the present study, all the rats first received preshocks in a Y maze. Then, half of the animals was immediately anaesthetized with halothane. The possible effects of halothane will be more easily detectable when there is no delay between preshocks and anaesthesht. One day after these pretreatments, all the animals were given an avoidance learning session in the Y maze. If the anaesthesia has aversive properties, then it should increase the impairment produced by the preshocks alone so that the performance of the preshocked-anaesthetized rats should be inferior to that of
C o p y r i g h t '~ 1979 Brain R e s e a r c h Publications Inc.~0031-9384/79/010025-05502.00/0
26
SCHMALTZ
the preshocked ones. In short, one should be able to detect the aversive effects of halothane anaesthesia, if any, by examining the performance of the animals during the avoidance conditioning session. METHOD Animals The animals were 85 male Sprague-Dawley rats weighing between 285-315 g at the start of the experiment. All the animals were housed individually, with ad lib food and water available in their home cages. They were handled once each morning during the week before the beginning of the experiment. Apparatus The apparatus used was a Y-nmze with an adjoining start box. All the walls were wooden. The 3 arms of the maze formed 120" angles, and each arm was 43 cm long, 12 cm wide and 30 cm high. The centrld alley was soperated from the start box by a ~ door, while each of the lateral arms had a slidlnll door 30 cm away from the ~ end. These doors ~ the e ~ t e r to isolate ~ animal in the goal box. The floor of the wmze, including the start box, consisted of parallel brass rods 3 nun in dian~ter lying at 10 m m intervals. This grid was connected to a constant current stimulator. The entire apparatus was covered with Perspex lids. Procedure The 85 animals were divided at random into 8 groups (N= 10, except for one Stoup with N=IS). For each animal, the experiment involved 2 sessions: pretreatment, then avoidance conditioning. Session 1: Pretreatment Non-anaesthetized groups. In 3 of the groups (M., M:j and M,o) the animals were placed individually in the maze where they remained for 30 win. They could not enter the start box, as door was kept closed. During this half hour the rats in these groups received, respectively, 0, 3 or 10 aversive preshocks (1,5 mA in pulsed train~--0.1 sec on; 0,4 sac oft). These l ~ k s (PS) were inesc,e p ~ and u n ~ e d , the onset, duration and intensity being controlled by the experimenter. The fact that the PS were uncontrollable considerably increased the aversiveness of the situation [10]. Immediately after the 30 rain spent in the maze, the aninmls were given sham anaesthesia (see below) then returned to their home cages. This sham a n a e s t h u was also given to rats in a control group Co, which were never placed in the maze, but simply renmined in their home cages. Anaesthetized groups. Groups C._j,, M._., M:,_,, and M,o-. received exactly the same pretreamumts (PS) as their counterparts described above, except that a FFdotex Mark 2 delivered a mixture of equal parts o f e l r and oxygen (flow 2 l/rain) with a 1,5% concentration of halothaae. The mixture passed through a Ptexigias box (25 cm×25 cm×25 cm), escaping through 4 small h ~ e s in the lid. Each rat was placed individually in this box for 60 rain. The average time to achieve narcosis was 40 sex. After spending an hour under the influence of anaesthetic, the unconscious animal was removed and placed in its home cage where it recovered fully
within 10 to 15 min. After waking there was no obvious sign of impairment in gross behaviour. The sham anaesthesia was performed in the same box, but there was no halothane in the air-oxygen mixture, and the time spent in the box was only 40 sec. Session 2: Avoidance Conditioning The active avoidance conditioning involved 15 trials in the Y-maze and took place one day after the first session. The same procedure, as previously described [9], was used for all the rats. The rat was placed in the start box at the beginning of the session. After 1 rain, the door was opened, allowing access to the maze alleys. Five sec later, the rat received footshock with the same characteristics as the pretreatment shock, except that now it could be controlled by the animal. The shock stopped when the animal, after reaching the end of one arm (first choice), turned round and went to the end of the other arm beyond the sliding door (second choice), This sliding door was then closed and the animal was left in the goal box for 30 sex before being placed in the start box for the I rain intertrial period. During the first trial it was thus necessary for the rat to cover the entire maze before escaping the aversive stimulus (escape response). During the following trials, the goal box (second choice on the first trial) was no longer electrified. The rat could now either make an escape response by entering the goal box after the shock had begun, or make a conditioned avoidance response (CAR) by entering the goal box directly--i.e., without making any spatial discrimination error--before the onset of the shock. Some animals, especially in the early stages of learning, did not escape the shock. In this case the experimenter intervened. During the first trial if, after a 20 sex stimulation period, the rat was still in the start box, it was pushed into the central alley and door was closed. If after 90 sex the animal had not made the escape response it was pushed into a goal box for 10 sex. Therefore shock could not last longer than I00 sex on any trial. The location of the goal box depended on the previous behaviour of the animal. The goal box was either the unexplored arm, or an arm was chosen at random if the rat had not explored either arm. During the remaining trials, the procedure was the same except that the goal box was already determined. RESULTS Behavioural Observations During Pretreatment While the rats were given these 3 uncontrollable PS, their most frequent response was flight. In some a n i ~ (8/20), freezing appeared on the third PS. Among those animals which were submitted to 10 uncontrollable PS, the same pattern was observed during the early shocks but freezing then increase in frequency. After the fifth PS, the animals froze consistently in response to the aversive stimulus. Avoidance Conditioning in Non-anaesthetized Groups During the active avoidance conditioning we recorded the duration of stimulation on each trial (range: 0-100 sec)for each rat and also the total duration of stimulation summed over all 15 trials. This enabled us to calculate, respectively, the mean duration of stimulation (MDS) per trial for a given group and the mean total duration of stimulation (MTDS) for the group. A Preshock×Anaesthesia analysis of variance
AVERSIVE EFFECTS OF HALOTHANE
27
demonstrated a statistically significant preshock effect, F(2,59)= 14.47, p
