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The Implication of Noradrenaline in Avoidance Learning in the Rat
The Implication of Noradrenaline in Avoidance Learning in the Rat K. D. CAIRNCROSS, SUE SCHOFIELD
AND
H. G . KING
School of Biological Sciences, Macquarie University, North Ryde, N.S. W. 2113 (Australia)
It has been reported that rats rendered anosmic following bilateral section of their olfactory tracts show a performance deficit in aversive learning (Marks et al., 1971; Cairncross and King, 1971). The anatomical and physiological consequences of olfactory deafferentation are profound, and may be enumerated as follows: ( i ) Degeneration occurs in primary olfactory neurones passing to the allocortex (in the terminology of Pigache, 1970). (ii) Following primary neurone degeneration, transneuronal degeneration occurs involving the pyramidal cells of the allocortex, which is characterised in the rat by a lack of dendritic proliferation (Jones and Thomas, 1962; White and Westrum, 1964). (iii) Following unilateral section of an olfactory tract, a reduction in telencephalic noradrenaline (NA) occurs in the side ipsilateral to the lesion, with no concomitant significant reduction in hypothalamic NA (Pohorecky et al., 1969). On the basis of the facts presented it appeared that the performance deficit in learning in aversive situations might be related to a reduction in NA availability in the telencephalon. Accordingly, male rats (Carworth CSF strain) 90 days old at the start of the experiment, were placed on a reverse 12-h night-day schedule, and housed in conditions of constant temperature and humidity. Following 14 days’ equilibration, the experimental group were subjected to surgery, consisting of section of the lateral olfactory tracts. Control animals underwent the same surgical procedure without olfactory tract section. The animals were sacrificed in groups of 6, 1-5 weeks after the surgical procedure. The brains were removed, and the cortex and hypothalamus assayed for endogenous NA after the spectrofluorimetric method of Haggendal (1963). The results of this experiment are illustrated in Fig. 1. It can be seen that the procedure produced a reduction in cortical NA 14 days after the completion of surgery, and that there was no recovery in cortical NA levels during the ensuing 3 weeks. In accord with the findings of Pohorecky et a]. (1969) there was no significant change in hypothalamic NA. It appeared, therefore, that bilateral olfactory tract section produced a reduction in endogenous NA specific to the cortical region. Once the specificity of the surgical procedure on cortical endogenous NA levels was established, animals rendered anosmic were subjected to behavioural testing 14-21 days following surgery. The behavioural method consisted of one-way avoidance References R . 484485
K. D. CAIRNCROSS et al.
482 y g NA/g
CORTEX
0.16
0.14
0.12
0.x
0.08
0.M
0
3
2
1
TIME (weeks)
4
Fig. 1. Reduction in cortical noradrenaline following bilateral olfactory tract section. 0.6
0.5
.
0.4
-
0.3
-
0.2
-
0.1
5 4
1
1-5
6-10
11-15
16-20
21-25
TRIALS IN BUXKS OF FIVE
Fig. 2. Avoidance responding on a 1-way task in anosmic and sham operated rats. A, anosmic; S, sham.
learning. The performance of the anosmic animals was compared with sham-operated controls. The results of this experiment are illustrated in Fig. 2. It can be seen that sham-operated animals quickly learned to escape the aversive stimulus, whereas the anosmic animals showed no such learning ability. It appeared,
483
NORADRENALINE IN AVOIDANCE LEARNING
therefore, that a correlation existed between the physiological and behavioural parameters examined. To test this hypothesis further, anosmic animals subjected to fear conditioning prior to the aversive avoidance task were examined. It was found that the anosmic group performed poorly when compared with the control group in such a situation. These observations further substantiated the hypothesis that a reduction in cortical NA availability reduced learning capacity. It was argued, therefore, that a pharmacological treatment aimed at increasing cortical NA availability should overcome the DAY 2
0 2
1.1
wa
n-a
DAY 6
'Im
W ~ I
I-10
TRIALS
TRIALS
a
a
c)
Fig. 3. Avoidance responding on a 1-way task following fear conditioning in anosmic rats. Group A received amitriptyline (1.5 mg/kg i.p., b.d.), group P received no drug, but a similar volume of isotonic saline i.p. w
NAh
CORTEX 014
-
-----_-__
@------
010
008
1
>\
//
\
0.04
0.02
/ /
o'w-
-
2
/
0.12 -
/ I
f--4'/
\
/
I
I
I
I
I
/
/
//
484
K. D. CAIRNCROSS
et al.
learning decrement exhibited by the anosmic animals. In this regard, the cortical and peripheral actions of the tricyclic anti-depressant drug amitriptyline were considered. This drug has been widely used in the treatment of depressive illness for many years, although its precise mode of action remains obscure. It is known, however, that amitriptyline and its metabolic derivatives enhance the peripheral actions of NA (Cairncross, 1965; Cairncross et al., 1967), and that this action relates to the ability of the drug to inhibit the re-uptake of NA into the nerve ending. (Story and Story, 1968). Similar activity has been demonstrated for amitriptyline in the central nervous system (Schubert et al., 1970; Cairncross and Martensz, 1972). Accordingly amitriptyline was administered to anosmic rats for a 14-day period. A control group of anosmic animals received a similar volume of isotonic saline. The animals were then tested in one-way avoidance learning, following prior fear conditioning. Immediately following behavioural testing, the animals were killed, their brains removed, and the cortex assayed for endogenous NA. The behavioural results obtained are presented in Fig. 3. Perusal of this figure illustrates that amitriptyline has a biphasic effect on behavioura1 performance. There is an initial reduction in performance in the amitriptylinetreated group apparent on day 2. By day 6 the amitriptyline-treated animals equate in performance with the placebo-treated anosmic animals, and by day 10 are statistically superior in performance to the placebo group, a situation remarkably similar to that described for the clinical situation (Hordern, 1965). The effects of amitriptyline on endogenous NA levels in the cortex are illustrated in Fig. 4. Again, a biphasic effect is evident. The initial response is a reduction in cortical NA, maximal at day 3, recovery to pre-drug anosmic levels by days 6-8, and thereafter an elevation of cortical endogenous NA levels to those found in non-operated control animals. The correlation between the improvement in behavioural performance and the recovery of endogenous NA to control values suggests that the supposition implicating NA in the development of avoidance learning is a phenomenon worthy of further investigation. It is suggested, also, that the results as presented offer a tentative explanation for the mode of action of amitriptyline in the clinical situation.
ACKNOWLEDGMENTS
The work was supported by Grant No. A65/15506 from the Australian Research Grants Committee.
REFERENCES
K. D. (1965) On the peripheral pharmacology of arnitriptyline.Arch. int. Pharrnacodyn., CAIRNCROSS,
154,43848.
CAIRNCROSS, K. D. AND KING, M. G . (1971) Facilitation of avoidance learning in anosrnic rats by amitriptyline. Proc. Aust. physiol. pharmacol. SOC.,2, 25.
NORADRENALINE IN AVOIDANCE LEARNING
485
CAIRNCROSS, K. D. AND MARTENSZ, N. D. (1972) Unpublished results. CAIRNCROSS, K. D., MCCULLOCH, MARIONW., STORY,D. F. AND TRINKER, F. (1967) Modification of synaptic transmission in the superior cervical ganglion by epinephrine, norepinephrine and nortriptyline. Int. J. Neuropharmacol., 6, 293-300. HAGGENDALL, .I.(1963) An improved method for fluorimetric determination of small amounts of adrenaline and noradrenaline in plasma and tissues. Acta physiol. scand., 59, 242-254. HORDERN, A. (1965) The antidepressant drugs. New Engl. J. Med., 272, 1159-1169. JONES,W. H. AND THOMAS, D. B. (1962) Changes in the dendritic organisation of neurones in the cerebral cortex following deafferentiation. J . Anaf. (Lond.), 96, 359-374. MARKS,H. E., REMLEY, W. R., SEAGO,J. D. AND HASTINGS, D. W. (1971) The effects of bilateral lesion of olfactory bulbs of rats on measures of learning and motivation. Physiol. Behav., 7 , 1-6. PIGACHE, R. M. (1970) The anatomy of the “paleocortex”, a critical review. In Reviews of Anatomy, Embryology and Cell Biology, Springer, Berlin, pp. 1-61. POHORECKY, L. A., ZIGMUND, M. J., REIMER, L. AND WURTMAN, R. G. (1969) Olfactory bulb removal --effects on brain norepinephrine. Proc. natl. Acad. Sci. (US.),62, 1052-1055. SCHUBERT, J., NYBACH, H. AND SEDVALL, G . C. (1970) Effect of antidepressant drugs on accumulation and disappearance of monoamines formed in vivo from labelled precursors in mouse brain. J. Pharm. Pharmacol., 22, 136-139. STORY, D. F. AND STORY,M. (1968) Studies on the uptake of H3-noradrenaline by isolated guinea-pig atrial tissue. Communication presented to Aust. Soc. elin. exp. Pharmacol., p. 38. WHITE,L. E. AND WESTRUM, L. E. (1964) Dendritic spine changes in prepyriform cortex following olfactory bulb lesions-rat, Golgi method. Anat. Rec., 148, 410-411.
AND
H. G . KING
School of Biological Sciences, Macquarie University, North Ryde, N.S. W. 2113 (Australia)
It has been reported that rats rendered anosmic following bilateral section of their olfactory tracts show a performance deficit in aversive learning (Marks et al., 1971; Cairncross and King, 1971). The anatomical and physiological consequences of olfactory deafferentation are profound, and may be enumerated as follows: ( i ) Degeneration occurs in primary olfactory neurones passing to the allocortex (in the terminology of Pigache, 1970). (ii) Following primary neurone degeneration, transneuronal degeneration occurs involving the pyramidal cells of the allocortex, which is characterised in the rat by a lack of dendritic proliferation (Jones and Thomas, 1962; White and Westrum, 1964). (iii) Following unilateral section of an olfactory tract, a reduction in telencephalic noradrenaline (NA) occurs in the side ipsilateral to the lesion, with no concomitant significant reduction in hypothalamic NA (Pohorecky et al., 1969). On the basis of the facts presented it appeared that the performance deficit in learning in aversive situations might be related to a reduction in NA availability in the telencephalon. Accordingly, male rats (Carworth CSF strain) 90 days old at the start of the experiment, were placed on a reverse 12-h night-day schedule, and housed in conditions of constant temperature and humidity. Following 14 days’ equilibration, the experimental group were subjected to surgery, consisting of section of the lateral olfactory tracts. Control animals underwent the same surgical procedure without olfactory tract section. The animals were sacrificed in groups of 6, 1-5 weeks after the surgical procedure. The brains were removed, and the cortex and hypothalamus assayed for endogenous NA after the spectrofluorimetric method of Haggendal (1963). The results of this experiment are illustrated in Fig. 1. It can be seen that the procedure produced a reduction in cortical NA 14 days after the completion of surgery, and that there was no recovery in cortical NA levels during the ensuing 3 weeks. In accord with the findings of Pohorecky et a]. (1969) there was no significant change in hypothalamic NA. It appeared, therefore, that bilateral olfactory tract section produced a reduction in endogenous NA specific to the cortical region. Once the specificity of the surgical procedure on cortical endogenous NA levels was established, animals rendered anosmic were subjected to behavioural testing 14-21 days following surgery. The behavioural method consisted of one-way avoidance References R . 484485
K. D. CAIRNCROSS et al.
482 y g NA/g
CORTEX
0.16
0.14
0.12
0.x
0.08
0.M
0
3
2
1
TIME (weeks)
4
Fig. 1. Reduction in cortical noradrenaline following bilateral olfactory tract section. 0.6
0.5
.
0.4
-
0.3
-
0.2
-
0.1
5 4
1
1-5
6-10
11-15
16-20
21-25
TRIALS IN BUXKS OF FIVE
Fig. 2. Avoidance responding on a 1-way task in anosmic and sham operated rats. A, anosmic; S, sham.
learning. The performance of the anosmic animals was compared with sham-operated controls. The results of this experiment are illustrated in Fig. 2. It can be seen that sham-operated animals quickly learned to escape the aversive stimulus, whereas the anosmic animals showed no such learning ability. It appeared,
483
NORADRENALINE IN AVOIDANCE LEARNING
therefore, that a correlation existed between the physiological and behavioural parameters examined. To test this hypothesis further, anosmic animals subjected to fear conditioning prior to the aversive avoidance task were examined. It was found that the anosmic group performed poorly when compared with the control group in such a situation. These observations further substantiated the hypothesis that a reduction in cortical NA availability reduced learning capacity. It was argued, therefore, that a pharmacological treatment aimed at increasing cortical NA availability should overcome the DAY 2
0 2
1.1
wa
n-a
DAY 6
'Im
W ~ I
I-10
TRIALS
TRIALS
a
a
c)
Fig. 3. Avoidance responding on a 1-way task following fear conditioning in anosmic rats. Group A received amitriptyline (1.5 mg/kg i.p., b.d.), group P received no drug, but a similar volume of isotonic saline i.p. w
NAh
CORTEX 014
-
-----_-__
@------
010
008
1
>\
//
\
0.04
0.02
/ /
o'w-
-
2
/
0.12 -
/ I
f--4'/
\
/
I
I
I
I
I
/
/
//
484
K. D. CAIRNCROSS
et al.
learning decrement exhibited by the anosmic animals. In this regard, the cortical and peripheral actions of the tricyclic anti-depressant drug amitriptyline were considered. This drug has been widely used in the treatment of depressive illness for many years, although its precise mode of action remains obscure. It is known, however, that amitriptyline and its metabolic derivatives enhance the peripheral actions of NA (Cairncross, 1965; Cairncross et al., 1967), and that this action relates to the ability of the drug to inhibit the re-uptake of NA into the nerve ending. (Story and Story, 1968). Similar activity has been demonstrated for amitriptyline in the central nervous system (Schubert et al., 1970; Cairncross and Martensz, 1972). Accordingly amitriptyline was administered to anosmic rats for a 14-day period. A control group of anosmic animals received a similar volume of isotonic saline. The animals were then tested in one-way avoidance learning, following prior fear conditioning. Immediately following behavioural testing, the animals were killed, their brains removed, and the cortex assayed for endogenous NA. The behavioural results obtained are presented in Fig. 3. Perusal of this figure illustrates that amitriptyline has a biphasic effect on behavioura1 performance. There is an initial reduction in performance in the amitriptylinetreated group apparent on day 2. By day 6 the amitriptyline-treated animals equate in performance with the placebo-treated anosmic animals, and by day 10 are statistically superior in performance to the placebo group, a situation remarkably similar to that described for the clinical situation (Hordern, 1965). The effects of amitriptyline on endogenous NA levels in the cortex are illustrated in Fig. 4. Again, a biphasic effect is evident. The initial response is a reduction in cortical NA, maximal at day 3, recovery to pre-drug anosmic levels by days 6-8, and thereafter an elevation of cortical endogenous NA levels to those found in non-operated control animals. The correlation between the improvement in behavioural performance and the recovery of endogenous NA to control values suggests that the supposition implicating NA in the development of avoidance learning is a phenomenon worthy of further investigation. It is suggested, also, that the results as presented offer a tentative explanation for the mode of action of amitriptyline in the clinical situation.
ACKNOWLEDGMENTS
The work was supported by Grant No. A65/15506 from the Australian Research Grants Committee.
REFERENCES
K. D. (1965) On the peripheral pharmacology of arnitriptyline.Arch. int. Pharrnacodyn., CAIRNCROSS,
154,43848.
CAIRNCROSS, K. D. AND KING, M. G . (1971) Facilitation of avoidance learning in anosrnic rats by amitriptyline. Proc. Aust. physiol. pharmacol. SOC.,2, 25.
NORADRENALINE IN AVOIDANCE LEARNING
485
CAIRNCROSS, K. D. AND MARTENSZ, N. D. (1972) Unpublished results. CAIRNCROSS, K. D., MCCULLOCH, MARIONW., STORY,D. F. AND TRINKER, F. (1967) Modification of synaptic transmission in the superior cervical ganglion by epinephrine, norepinephrine and nortriptyline. Int. J. Neuropharmacol., 6, 293-300. HAGGENDALL, .I.(1963) An improved method for fluorimetric determination of small amounts of adrenaline and noradrenaline in plasma and tissues. Acta physiol. scand., 59, 242-254. HORDERN, A. (1965) The antidepressant drugs. New Engl. J. Med., 272, 1159-1169. JONES,W. H. AND THOMAS, D. B. (1962) Changes in the dendritic organisation of neurones in the cerebral cortex following deafferentiation. J . Anaf. (Lond.), 96, 359-374. MARKS,H. E., REMLEY, W. R., SEAGO,J. D. AND HASTINGS, D. W. (1971) The effects of bilateral lesion of olfactory bulbs of rats on measures of learning and motivation. Physiol. Behav., 7 , 1-6. PIGACHE, R. M. (1970) The anatomy of the “paleocortex”, a critical review. In Reviews of Anatomy, Embryology and Cell Biology, Springer, Berlin, pp. 1-61. POHORECKY, L. A., ZIGMUND, M. J., REIMER, L. AND WURTMAN, R. G. (1969) Olfactory bulb removal --effects on brain norepinephrine. Proc. natl. Acad. Sci. (US.),62, 1052-1055. SCHUBERT, J., NYBACH, H. AND SEDVALL, G . C. (1970) Effect of antidepressant drugs on accumulation and disappearance of monoamines formed in vivo from labelled precursors in mouse brain. J. Pharm. Pharmacol., 22, 136-139. STORY, D. F. AND STORY,M. (1968) Studies on the uptake of H3-noradrenaline by isolated guinea-pig atrial tissue. Communication presented to Aust. Soc. elin. exp. Pharmacol., p. 38. WHITE,L. E. AND WESTRUM, L. E. (1964) Dendritic spine changes in prepyriform cortex following olfactory bulb lesions-rat, Golgi method. Anat. Rec., 148, 410-411.