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Reflex habituation and potentiation in Rana pipiens
REFLEX HABITUATION AND POTENTIATION IN RANA PIPIENS BY
DANIEL P. KIMBLE AND ROBERTA SHOCKLEY RAY Department of Psychology, University of Oregon .
When an initially arousing stimulus is repeatedly presented to an animal with no accompanying reinforcement, a gradual diminution of the response typically occurs . This is habituation, and is a well known and important behavioural phenomenon (John, 1961) . Recently, however, Franzisket (1963) has reported that a series of nonreinforced presentations of a stimulus led not to habituation, but to an increase in the rate of responding . Specifically, he reported that the daily number of wiping reflexes elicited from both spinal and intact frogs (Rana esculenta) increased from 4 to 14 per cent . to 60 to 90 per cent. over a 5 to 7 day period. These reflex movements were elicited by stimulating the back skin of the frogs with a von Frey bristle. He interpreted these and similar results as being the result of training and invoked two neurophysiological concepts to explain his results. These concepts were : (1) Facilitation of the underlying synapses "probably identical with the post-tetanic potentiation as described by D . P . C. Lloyd (1949) and by Eccles & McIntyre (1951)" . (2) A longer lasting "hypertrophy ("Aktivitatshypertrophie") Verworn (1910) of the coordinating neurons of the reflex centre" . Since an increase in response rate to repeated nonreinforced stimuli is in contradiction to more typical findings, including other work with amphibians (Kuczka, 1956), it was decided to attempt a replication of Franzisket's study, and also to determine if the details of the stimulus presentation might be critical for the resulting response rate .
same as that used by Franzisket (1963) . In order to prevent the escape of the frogs during the trial periods, stimulation was done under red light . This technique effectively reduces the usual amount of movement, making blinding or other undesirable physical intervention unnecessary . These procedures were also the same as those employed by Franzisket (1963) . Franzisket reported that there are 3 reflexogenous zones on the dorsal skin of the frog . Stimulation of the anterior zone produces a wiping movement of the arm such that the front foot passes over the head from occiput to nose . Stimulation of the caudal zone produces a wiping movement of the heel of the ipsilateral hind leg . A stimulus applied to the major reflexogenous zone in the middle portion of the back elicits a wiping movement by the hind foot over the back. It was this last mentioned reflex movement which was studied in the present experiment as well as in Franzisket's report . Franzisket reported potentiation of the wiping reflex as a result of repeatedly eliciting the reflex . However, as he points out, this reflex can be obtained from a rather general area or reflexogenous zone . We decided to carefully delimit and maintain the precise locus of stimulation within this major reflexogenous zone in one group of animals, to determine if habituation could be obtained in this situation by increasing the similarity of the repeated stimulations . Therefore, for the Ss in Group A (reflex potentiation group), Franzisket's procedures were followed as exactly as could be determined from his report. The stimulus was applied within the right half of the major dorsal reflexogenous area from which the foot wiping reflex is characteristically obtained . Care was taken to apply the stimulus within this area, and thus repeatedly elicit the same reflex movement . In Group B (reflex habituation group), the same procedures were followed with one important addition . For these Ss, a precise spot within the right half of the major reflexogenous zone was determined and maintained on the basis of the skin markings of that particular S. This specific stimulation spot was repeatedly stimulated for that S, although the point of stimulation varied slightly from S to S within
Method Subjects The Ss were 12 frogs (Rana pipiens) obtained commercially. They were maintained throughout the experiment in separate cage compartments which simulated a natural environment . Test Procedure The Ss were randomly assigned to one of two groups . Thus, the N in each group was 6 . All Ss were stimulated with a standardized bristle, No . 10 in the von Frey series, having a radius of 0 .0925 mm . and a force of 0 . 8 g. This was the 530
KIMBLE & RAY : REFLEX HABITUATION IN RANA PIPIENS
Group B . In both groups, the stimulation was applied 100 times per day at 10 second intervals for a total of 12 days . Thus, not only was the same reflex repeatedly elicited, as in Group A, but also the stimuli were as nearly identical as possible . Results and Discussion The number of wiping reflexes elicited during each daily 100 trials session for each S is given in Table I . The mean and median responses are also given for each group for each day . As can be seen in Table I, the number of daily responses increased steadily throughout the experiment for the Ss in Group A, while clear cut response diminution or habituation occured in Ss from Group B . Fig. 1 presents these data in graphic form . Although no differences appeared between the two experimental groups on the first training day, significant differences soon developed . By the sixth experimental day, there was no overlap in the scores for the Ss in the two groups. A sign test (Dixon & Massey, 1957) was performed, comparing the data of Day 1 with the data of Day 12 for each group . All Ss in Group A (reflex potentiation) had higher scores for Day 12 than they had for Day. I . Just the reverse was true for the Ss in Group B (reflex habituation) . These differences were significant at <0 .05 level of probability .
531
These differences appear to be due to the difference in the method of stimulus presentation . The results of the Group B Ss indicate that if the locus of stimulation is carefully controlled so that the series of stimuli are more truly "repeated" stimuli, a gradual diminution, or habituation of responding does occur . However, the Ss in Group A, as well as those of Franzisket (1963) indicate that any slight variation from stimulation to stimulation, even though the same reflex movement is obtained, can lead to an increase rather than a decrease in the rate of responding . Habituation in these Ss seems to depend on not only eliciting the same response with repeated nonreinforced stimulation, but upon repeating the nature and locus of the stimulation as closely as possible . Although habituation of the wiping reflex was obtained by holding the locus of stimulation as constant as possible, reflex potentiation as reported by Franzisket was also found in the present experiment . The concepts of induced neuronal hypertrophy and post-tetanic potentiation have been derived primarily from findings based on particular synaptic junctions, usually monosynaptic reflex centres (Eccles, 1961), and very intensive stimulation (i .e . 555 stimuli/ second for 12 seconds), quite unlike the stimulation used in the present reports . The reflex potentiation found in our study occurs when the
Table I. Number of Wiping Reflexes Elicited per 100 Stimuli .
Group A (Reflex potentiation) Day A-1 A-2 A-3 A-4 A-5 A-6 Mean Median
1
2
3
4
5
6
36 18 30 41 15 1 47
41 26 35 36 23 51
35 22 40 47 32 45
52 30 38 52 26 63
46 32 47 57 23 67
31 .2
35 . 3
36 . 8
43 . 5
33 .0
35 . 5
37 . 5
1 1
i
7
8
9
64 28 49 63 31 60
56 30 48 65 33 58
62 33 53 63 30 62
45 . 3
49 . 1
48 . 3
45 . 0
46 . 5
54 . 5
34 37 37 33 12 35
26 33 43 29 13 24
18 28 30 23 16 20
32
31 . 3
28
32 . 5
34 . 5
27 . 5
Group B (Reflex habituation) I B-1 32 30 B-2 41 35 B-3 46 41 B-4 37 39 B-5 14 17 B-6 28 30 Mean
33
l
10
11
12
67 29 50 67 28 60
64 35 56 61 36 64
70 33 54 70 35 61
68 34 60 71 38 64
50 . 5
50 . 2
52 . 7
53 . 8
55 . 8
52 . 0
57 . 5
55 . 0
58 . 5
57 . 0
62 . 0
22 30 30 25 8 22
21 28 28 24 12 20
18 25 23 26 10 18
18 30 24 22 8 19
16 22 26 19 10 15
18 23 21 17 11 20
14 20 20 18 8 17
22 .5
22 . 8
22 .2
20 .0
20 .2
18 .0
18 . 5
16 .2
21 .5
23 . 5
22 . 5
20•5
20 . 5
17 . 5
19 .0
17 . 5
1
1
i Median
34 . 5
ANIMAL BEHAVIOUR, XIII, 4
532 J D IN
0
REFLEX POTENTIATION GROUP 60
• • 50 X w J LL 40
a LL
0 aw 30 m z 20 z
REFLEX HABITUATION GROUP
• W 10
I 56789101112 DAY Fig. 1 . Median number of daily wiping reflex movements per 100 stimuli elicited in reflex potentiation and reflex habituation Ss. locus of stimulation varies slightly within the for the elicitation of the wiping reflex within the reflexogenous zone, thus presumably activating reflexogenous zone . This possibility is in fact a more diffuse neural substratum than that used strengthened by Franzisket's report of lowered in the post-tetanic potentiation work . Therefore, threshold for eliciting the wiping reflex . This we do not feel that the concepts of induced lowering of the threshold corresponded in time neuronal atrophy or post-tetanic potentiation with the increase in response rate. Franzisket need necessarily be involved to account for this found that in an untrained frog a pressure of increase in response rate . It is not possible, 24 to 27 g./mm. 2 was necessary to obtain the first however, to completely rule out this type of reflex, but that after 5 to 7 days of training a synaptic modification as a possible explanation previously subliminal stimulation of only 9 to of this phenomenon . 12 g ./mm .2 was sufficient to elicit the first reflex An alternative explanation which appears to for that day. be functionally equivalent but which involves Thus, we would suggest that subliminal excitdifferent neural modifications can be offered . ation of adjacent areas within the reflexogenous Stimulation which varies slightly in locus from zone may account for the observed response trial to trial may not only elicit the wiping reflex potentiation . When these adjacent areas are some percentage of the time, but might also exminimally involved by holding the locus of stimert a sustained but subliminal effect on adjacent ulation constant (as in Group B), habituation to area so as to slightly raise its excitability . A the stimulation will occur, indicating a relative stimulus presented more directly to this adjacent specificity to the stimulus for the phenomenon area might then elicit a reflex more readily than of reflex habituation . would be the case without the previous subliminal Summary excitation . Reflex potentiation would then occur as these subliminal effects lowered the threshold This study was an attempt to further elucidate
KIMBLE & RAY : REFLEX HABITUATION IN RANA PIPIENS
the nature of habituation . Two groups of frogs (Rana pipiens) were stimulated 100 times daily for 12 days with a No . 10 von Frey bristle . In one group, the locus of stimulation was maintained within the general area which elicits the wiping reflex of the ipsilateral hind foot . In the other group, the same reflex movement was elicited from a carefully delimited spot within the reflexogenous zone which was maintained from day to day on the basis of skin markings . In the former group the rate of responding increased from day to day, while in the latter group, habituation to the repeated stimulation occurred . These results were interpreted to mean that habituation depends upon not only eliciting the same response with a series of repeated, nonreinforced stimuli, but that it is necessary as well to insure that the nature and location of the stimulus is repeated as closely as possible. Acknowledgements The authors would like to acknowledge the assistance of Malcolm Ray throughout the ex-
533
periment . The experiment was supported by funds from United States Public Health Grant MH 08545-01 . REFERENCES Dixon, W . J . & Massey, F . J . (1957) . Introduction to Statistical Analysis . New York : McGraw-Hill . Eccles, J . C. (1961). The effects of use and disuse on synaptic function . In, J . F. Delafresnaye (Ed.), Brain Mechanisms and Learning. Oxford : Blackwell Scientific Publications, Pp. 335-352 . Eccles, J. C . & McIntyre, A. K. (1951). Plasticity of mammalian monosynaptic reflexes . Nature, 167, 466 .
Franzisket, L. (1963) . Characteristics of instinctive behavior and learning in reflex activity of the frog . Anim. Behav., 11, 318 .
John, E. R . (1961) . High nervous functions : brain functions and learning . Ann. Rev. Physiol., 23, 451 . Kuczka, H . (1956) . Verhaltenphysiologische Untersuchungen uber die Wischhandlung der Erdkrote . (Bufo bufo L.).
Z.
Tierpsychol., 13, 185 .
Lloyd, D . P . C. (1949) . Post-tetanic potentiation of response in monosynaptic reflex pathway of the spinal cord . J. Gen . Physiol., 33, 147. (Accepted for publication 19th November, 1964 ; Ms. number : 519).
DANIEL P. KIMBLE AND ROBERTA SHOCKLEY RAY Department of Psychology, University of Oregon .
When an initially arousing stimulus is repeatedly presented to an animal with no accompanying reinforcement, a gradual diminution of the response typically occurs . This is habituation, and is a well known and important behavioural phenomenon (John, 1961) . Recently, however, Franzisket (1963) has reported that a series of nonreinforced presentations of a stimulus led not to habituation, but to an increase in the rate of responding . Specifically, he reported that the daily number of wiping reflexes elicited from both spinal and intact frogs (Rana esculenta) increased from 4 to 14 per cent . to 60 to 90 per cent. over a 5 to 7 day period. These reflex movements were elicited by stimulating the back skin of the frogs with a von Frey bristle. He interpreted these and similar results as being the result of training and invoked two neurophysiological concepts to explain his results. These concepts were : (1) Facilitation of the underlying synapses "probably identical with the post-tetanic potentiation as described by D . P . C. Lloyd (1949) and by Eccles & McIntyre (1951)" . (2) A longer lasting "hypertrophy ("Aktivitatshypertrophie") Verworn (1910) of the coordinating neurons of the reflex centre" . Since an increase in response rate to repeated nonreinforced stimuli is in contradiction to more typical findings, including other work with amphibians (Kuczka, 1956), it was decided to attempt a replication of Franzisket's study, and also to determine if the details of the stimulus presentation might be critical for the resulting response rate .
same as that used by Franzisket (1963) . In order to prevent the escape of the frogs during the trial periods, stimulation was done under red light . This technique effectively reduces the usual amount of movement, making blinding or other undesirable physical intervention unnecessary . These procedures were also the same as those employed by Franzisket (1963) . Franzisket reported that there are 3 reflexogenous zones on the dorsal skin of the frog . Stimulation of the anterior zone produces a wiping movement of the arm such that the front foot passes over the head from occiput to nose . Stimulation of the caudal zone produces a wiping movement of the heel of the ipsilateral hind leg . A stimulus applied to the major reflexogenous zone in the middle portion of the back elicits a wiping movement by the hind foot over the back. It was this last mentioned reflex movement which was studied in the present experiment as well as in Franzisket's report . Franzisket reported potentiation of the wiping reflex as a result of repeatedly eliciting the reflex . However, as he points out, this reflex can be obtained from a rather general area or reflexogenous zone . We decided to carefully delimit and maintain the precise locus of stimulation within this major reflexogenous zone in one group of animals, to determine if habituation could be obtained in this situation by increasing the similarity of the repeated stimulations . Therefore, for the Ss in Group A (reflex potentiation group), Franzisket's procedures were followed as exactly as could be determined from his report. The stimulus was applied within the right half of the major dorsal reflexogenous area from which the foot wiping reflex is characteristically obtained . Care was taken to apply the stimulus within this area, and thus repeatedly elicit the same reflex movement . In Group B (reflex habituation group), the same procedures were followed with one important addition . For these Ss, a precise spot within the right half of the major reflexogenous zone was determined and maintained on the basis of the skin markings of that particular S. This specific stimulation spot was repeatedly stimulated for that S, although the point of stimulation varied slightly from S to S within
Method Subjects The Ss were 12 frogs (Rana pipiens) obtained commercially. They were maintained throughout the experiment in separate cage compartments which simulated a natural environment . Test Procedure The Ss were randomly assigned to one of two groups . Thus, the N in each group was 6 . All Ss were stimulated with a standardized bristle, No . 10 in the von Frey series, having a radius of 0 .0925 mm . and a force of 0 . 8 g. This was the 530
KIMBLE & RAY : REFLEX HABITUATION IN RANA PIPIENS
Group B . In both groups, the stimulation was applied 100 times per day at 10 second intervals for a total of 12 days . Thus, not only was the same reflex repeatedly elicited, as in Group A, but also the stimuli were as nearly identical as possible . Results and Discussion The number of wiping reflexes elicited during each daily 100 trials session for each S is given in Table I . The mean and median responses are also given for each group for each day . As can be seen in Table I, the number of daily responses increased steadily throughout the experiment for the Ss in Group A, while clear cut response diminution or habituation occured in Ss from Group B . Fig. 1 presents these data in graphic form . Although no differences appeared between the two experimental groups on the first training day, significant differences soon developed . By the sixth experimental day, there was no overlap in the scores for the Ss in the two groups. A sign test (Dixon & Massey, 1957) was performed, comparing the data of Day 1 with the data of Day 12 for each group . All Ss in Group A (reflex potentiation) had higher scores for Day 12 than they had for Day. I . Just the reverse was true for the Ss in Group B (reflex habituation) . These differences were significant at <0 .05 level of probability .
531
These differences appear to be due to the difference in the method of stimulus presentation . The results of the Group B Ss indicate that if the locus of stimulation is carefully controlled so that the series of stimuli are more truly "repeated" stimuli, a gradual diminution, or habituation of responding does occur . However, the Ss in Group A, as well as those of Franzisket (1963) indicate that any slight variation from stimulation to stimulation, even though the same reflex movement is obtained, can lead to an increase rather than a decrease in the rate of responding . Habituation in these Ss seems to depend on not only eliciting the same response with repeated nonreinforced stimulation, but upon repeating the nature and locus of the stimulation as closely as possible . Although habituation of the wiping reflex was obtained by holding the locus of stimulation as constant as possible, reflex potentiation as reported by Franzisket was also found in the present experiment . The concepts of induced neuronal hypertrophy and post-tetanic potentiation have been derived primarily from findings based on particular synaptic junctions, usually monosynaptic reflex centres (Eccles, 1961), and very intensive stimulation (i .e . 555 stimuli/ second for 12 seconds), quite unlike the stimulation used in the present reports . The reflex potentiation found in our study occurs when the
Table I. Number of Wiping Reflexes Elicited per 100 Stimuli .
Group A (Reflex potentiation) Day A-1 A-2 A-3 A-4 A-5 A-6 Mean Median
1
2
3
4
5
6
36 18 30 41 15 1 47
41 26 35 36 23 51
35 22 40 47 32 45
52 30 38 52 26 63
46 32 47 57 23 67
31 .2
35 . 3
36 . 8
43 . 5
33 .0
35 . 5
37 . 5
1 1
i
7
8
9
64 28 49 63 31 60
56 30 48 65 33 58
62 33 53 63 30 62
45 . 3
49 . 1
48 . 3
45 . 0
46 . 5
54 . 5
34 37 37 33 12 35
26 33 43 29 13 24
18 28 30 23 16 20
32
31 . 3
28
32 . 5
34 . 5
27 . 5
Group B (Reflex habituation) I B-1 32 30 B-2 41 35 B-3 46 41 B-4 37 39 B-5 14 17 B-6 28 30 Mean
33
l
10
11
12
67 29 50 67 28 60
64 35 56 61 36 64
70 33 54 70 35 61
68 34 60 71 38 64
50 . 5
50 . 2
52 . 7
53 . 8
55 . 8
52 . 0
57 . 5
55 . 0
58 . 5
57 . 0
62 . 0
22 30 30 25 8 22
21 28 28 24 12 20
18 25 23 26 10 18
18 30 24 22 8 19
16 22 26 19 10 15
18 23 21 17 11 20
14 20 20 18 8 17
22 .5
22 . 8
22 .2
20 .0
20 .2
18 .0
18 . 5
16 .2
21 .5
23 . 5
22 . 5
20•5
20 . 5
17 . 5
19 .0
17 . 5
1
1
i Median
34 . 5
ANIMAL BEHAVIOUR, XIII, 4
532 J D IN
0
REFLEX POTENTIATION GROUP 60
• • 50 X w J LL 40
a LL
0 aw 30 m z 20 z
REFLEX HABITUATION GROUP
• W 10
I 56789101112 DAY Fig. 1 . Median number of daily wiping reflex movements per 100 stimuli elicited in reflex potentiation and reflex habituation Ss. locus of stimulation varies slightly within the for the elicitation of the wiping reflex within the reflexogenous zone, thus presumably activating reflexogenous zone . This possibility is in fact a more diffuse neural substratum than that used strengthened by Franzisket's report of lowered in the post-tetanic potentiation work . Therefore, threshold for eliciting the wiping reflex . This we do not feel that the concepts of induced lowering of the threshold corresponded in time neuronal atrophy or post-tetanic potentiation with the increase in response rate. Franzisket need necessarily be involved to account for this found that in an untrained frog a pressure of increase in response rate . It is not possible, 24 to 27 g./mm. 2 was necessary to obtain the first however, to completely rule out this type of reflex, but that after 5 to 7 days of training a synaptic modification as a possible explanation previously subliminal stimulation of only 9 to of this phenomenon . 12 g ./mm .2 was sufficient to elicit the first reflex An alternative explanation which appears to for that day. be functionally equivalent but which involves Thus, we would suggest that subliminal excitdifferent neural modifications can be offered . ation of adjacent areas within the reflexogenous Stimulation which varies slightly in locus from zone may account for the observed response trial to trial may not only elicit the wiping reflex potentiation . When these adjacent areas are some percentage of the time, but might also exminimally involved by holding the locus of stimert a sustained but subliminal effect on adjacent ulation constant (as in Group B), habituation to area so as to slightly raise its excitability . A the stimulation will occur, indicating a relative stimulus presented more directly to this adjacent specificity to the stimulus for the phenomenon area might then elicit a reflex more readily than of reflex habituation . would be the case without the previous subliminal Summary excitation . Reflex potentiation would then occur as these subliminal effects lowered the threshold This study was an attempt to further elucidate
KIMBLE & RAY : REFLEX HABITUATION IN RANA PIPIENS
the nature of habituation . Two groups of frogs (Rana pipiens) were stimulated 100 times daily for 12 days with a No . 10 von Frey bristle . In one group, the locus of stimulation was maintained within the general area which elicits the wiping reflex of the ipsilateral hind foot . In the other group, the same reflex movement was elicited from a carefully delimited spot within the reflexogenous zone which was maintained from day to day on the basis of skin markings . In the former group the rate of responding increased from day to day, while in the latter group, habituation to the repeated stimulation occurred . These results were interpreted to mean that habituation depends upon not only eliciting the same response with a series of repeated, nonreinforced stimuli, but that it is necessary as well to insure that the nature and location of the stimulus is repeated as closely as possible. Acknowledgements The authors would like to acknowledge the assistance of Malcolm Ray throughout the ex-
533
periment . The experiment was supported by funds from United States Public Health Grant MH 08545-01 . REFERENCES Dixon, W . J . & Massey, F . J . (1957) . Introduction to Statistical Analysis . New York : McGraw-Hill . Eccles, J . C. (1961). The effects of use and disuse on synaptic function . In, J . F. Delafresnaye (Ed.), Brain Mechanisms and Learning. Oxford : Blackwell Scientific Publications, Pp. 335-352 . Eccles, J. C . & McIntyre, A. K. (1951). Plasticity of mammalian monosynaptic reflexes . Nature, 167, 466 .
Franzisket, L. (1963) . Characteristics of instinctive behavior and learning in reflex activity of the frog . Anim. Behav., 11, 318 .
John, E. R . (1961) . High nervous functions : brain functions and learning . Ann. Rev. Physiol., 23, 451 . Kuczka, H . (1956) . Verhaltenphysiologische Untersuchungen uber die Wischhandlung der Erdkrote . (Bufo bufo L.).
Z.
Tierpsychol., 13, 185 .
Lloyd, D . P . C. (1949) . Post-tetanic potentiation of response in monosynaptic reflex pathway of the spinal cord . J. Gen . Physiol., 33, 147. (Accepted for publication 19th November, 1964 ; Ms. number : 519).