Psychological Reports, 1975, 37, 487-494. @ Psychological Reports 1975
WATER BALANCE A N D TITRATED PAIN THRESHOLDS W. E. FLYNN, C. A. SCHAUER, AND W. H. TEDFORD, JR. Southern Methodist University Summary.-Changes in sensitivity to electric foot shock were manipulated by subjecting 2 rats to 3 levels of water deprivation. Shock intensity was continuously titrated every 2 sec. through 20 increasing steps of alternating current. A bar-press reset the shock to its minimal value. The animals performed under conditions of 0, 21.5, and 45.5 hr. water deprivation. When water deprived, animals showed decrewed sensitivity to shock. Frequency of bar-press responses at each of the 20 shock values served as the dependent measure. Findings were discussed in terms of competing techniques for measurement of analgesia.
Geller and Axelrod (1968) have recently evaluated the use of operant techniques in the study of analgesics. They found reflexive measures (such as the tail flick response) to show high inter-experimenter variability and to be sensitive to compounds which are not usually clinically effective in pain reduction. That is, control animals injected with water, propandiol or saline showed more pain threshold elevation than animals under most dose levels of morphine. After rejecting avoidance and escape-avoidance techniques of pain measurement for similar reasons, they recommended the titration method of Weiss and Laties (1958) as the most efficient. The titration method (or fractional-escape as it is sometimes called) produces an increasing intensity of electric footshock to an animal. Each bar-press the animal makes, however, reduces the shock intensity one increment. Thus, animals "hold" the shock intensity at some threshold level. An alternate method is to have the response reset the shock level to zero. Recently, Houser and VanHart ( 1973) have adopted a shuttle-box escape technique for the evaluation of analgesics. Unfortunately, earlier work has shown pain thresholds derived from this technique to be sensitive to such clinically unresponsive manipulations as water deprivation (Misanin & Campbell, 1969). Reflexive measures of pain (vigorous movements, vocalizations or excessive urination) also appear to be sensitive to negative water loads (Griffiths, 1962). The present study was undertaken to examine the sensitivity of the titration technique to manipulations of water balance.
METHOD Two experimentally naive, male, Sprague-Dawley rats were used as Ss. They were 170 days old at the beginning of the study. Testing occurred in a Lehigh Valley small animal test cage which was inside a sound-insulated, environmental cubicle. White noise was continuously present.
488
W. E. FLYNN, ET AL.
The electrical stimulus was produced by a Foringer constant current shock generator with scrambler. During a portion of the procedure used for scaling, the output of the stimulator varied from 0.27 to 0.91 mA. For the remainder of the experiment, the output range was from 0.55 to 1.88 mA. During both settings, the stimulus was varied in 20 discrete and nearly equal steps. Pulses delivered to a stepping switch raised the intensity of the shock by one step every 2 sec. Whenever the lever in the experimental chamber was depressed, the output of the stimulator was reset to its minimal value. Output from the bar was fed through a pulseformer, so that the bar had to be released before a new response could be made. Counters connected to the stepping device recorded the total number of responses made at each step during 2-hr. sessions. During preuaining, Ss were housed in individual cages in a room illuminated on a 12-hr. dark, 12-hr. light cycle. The temperature and humidity were approximately constant. Throughout the experiment, Ss were allowed ad libitam access to Purina Laboratory Chow Checkers. Ss were allowed 30-min. daily access to water. More than 2 mo. of pretraining with 30-min. access to water stabilized daily body weights and water inmke before the experiment began. Ss were shaped to bar-press by presenting a .5-mA shock which was eliminated for 45 sec. by a response. After this response was shaped into a bar-press with a latency of less than 45 sec. from shock onset, 2 hr. daily titration sessions were initiated. Sessions were conducted 7 days per week, 2 hr. per day. The purpose of the first 2 stages of the experiment was to examine the measure of shock sensitivity. Both stages were conducted with Ss 0-ht. water deprived, that is, their 30-min. access to water occurred immediately prior to the 2-hr. experimental session. Ss were tested at a low shock range (0.27 to 0.91 mA) for 16 days. During this period, their performance stabilized. For the second stage of the scaling process, and for the remainder of the experiment, the output of the shock generator was set on the higher ranges (0.55 to 1.88 mA). Measures were taken at this stage for 22 days, during which time performance again stabilized. This latter stage also served as the 0-hr. deprived stage. The procedure for the 21.5-hr. deprived stage was identical to the previous 2 stages except for the watering schedule. During this stage, Ss received water immediately after each day's experimental session. In this way, Ss were 21.5-hr. deprived when the next session started. Data were collected for 11 days under this condition. For the following 24 days, Ss received water every other day immediately after an experimental session. A measure of the aversiveness of the shock was taken, however, every day. This gave 12 additional measures of aversiveness at 2 1.5-hr. deprivation, interspersed with 12 measures of aversiveness at 45.5-hr. deprivation. The final condition was a reinstatement of an earlier stage. For these 10
WATER BALANCE AND TITRATED PAIN THRESHOLDS
489
days, the animals' performance was measured immediately after having access to water.
RESULTS For each S the number of responses made at each of the 20 steps of increasing shock intensity was recorded for every experimental session. The measure of shock sensitivity used was a ratio of the number of responses made in the first 10 steps divided by the number of responses made in the last 10 steps. Thus, higher ratios indicate greater sensitivity to shock. Sensitivity ratios for the last 5 days in each treatment condition are shown in Fig. 1 for both Ss. Inspection of the figure suggests two conclusions. First, the sensitivity ratio appears to be responsive to increases in shock intensity. During the scaling procedure (Panels A and B) , this value increased when the electrical stimulus was changed from the low to the high shock range. Secondly, Ss' sensitivity to shock decreased when they were water deprived as indicated by B and E vs C and D. An analysis of variance performed on these dam confirmed a significant over-all treatment effect for each S (+ < .05). A 5 .
4 .
I
-
8
P
D
C
E
a1.5h..
N
Y : , 1
A
45.5 hr.
I . LO SHOCK H I SMOCK
0
0
H I SHOCK
11.5
HI
SHOCK
21.5 m d 45.5
H I SHOCX
0
HOURS OF WATER DEPRIVATION
FIG. 1. Percentage of Ss' responses that terminated low rather than high shock intensities. Panels B through D show effect of 0-, 21.5- and 45.5-hr. water deprivation. During the 45.5-hr. deprivation period, shack sensitivity was also determined on preceding days (Panel D, 21.5 hr.). The first panel ( A ) shows, for comparison, a lower range of shock output from the source.
W.
490
E. FLYNN, ET AL.
A Newman-Keuls contrast procedure was used to examine individual differences between the various experimental conditions. The scaling procedure which compared low vs high ranges of shock (Panel A vs B ) showed significant differences in the expected direction. That is, Ss responded more often on lower shock steps to the more intensive shock range. For both Ss, the difference between 0-hr. deprivation (Panel B) and both 21.5-hr. (Panel C ) and 45.5-hr. deprivation (Panel D ) proved to be significant (p .05). When Ss were deprived of water for 45.5 hr., however, sensitivity to shock was not significantly different (p > .05) from 21.5-hr. deprivation (Panel C vs D ) . Within the water-every-other-day period (Panel D) sensitivity was significantly lower during the 45.5-hr. deprived segment than during the 21.5-hr. deprived segment ( p < .05). A significant difference ( p < .05) was observed for both Ss during the 21.5-hr. deprived condition (Panel C ) compared to 21.5-hr. deprivation during the alternating 21.5- and 45.5-hr. deprived condition (Panel D) . N o significant differences ( p > .O5) were found between the second 0-hr. deprived condition and the return to that condition (Panel B vs E ) . Thus, manipulations of the independent variable of water deprivation did not appear to affect Ss' original reaction to shock at 0-hr. deprivation. Any fluctuations from the baseline value shown in Panels B and E which might be observed in the remaining panels should, therefore, represent some manipulation of water balance and not a simple effect of trials or learning. It should be noted thac water balance in the present study modified pain thresholds, but in some non-monotonic manner. For example, the 45.5-hr. deprivation condition failed to differ significantly from 21.5 hr. in Panel D. Moreover, during 21.5-hr. water deprivation, sensitivity was lower when water was offered daily than when water was given every 2 days. These discrepancies should be viewed in light of compensatory drinking behaviors observed during the 48-hr. deprivation period (see Fig. 2 ) . That is, animals apparently learned to drink and store more water under 45.5-hr. deprivation. Thus, water deficits in the 45.5-hr. condition may have been no different from the 21.5-hr. condition. Bolles (1967) has suggested thac rats will "learn to drink" much as they have been shown to learn to eat when deprived of food (Hebb, 1949; Reid & Finger, 1955). This means that the animal would learn to compensate for any existing deprivation state by overdrinking during those periods when water was made available. Fig. 2 gives the milliliters of water consumed and the body weights during the last 5 days of each condition for both Ss. An analysis of variance performed on these data indicated a significant treatment effect over conditions for both Ss ( p < .05). A Newman-Keuls contrast procedure showed thac Ss consumed significantly more water ( p .O5) when allowed access to water only on every other day (Panel D ) as opposed to every day (Panels A, B, C, and
WATER BALANCE A N D TITRATED PAIN THRESHOLDS W. E. FLYNN, C. A. SCHAUER, AND W. H. TEDFORD, JR. Southern Methodist University Summary.-Changes in sensitivity to electric foot shock were manipulated by subjecting 2 rats to 3 levels of water deprivation. Shock intensity was continuously titrated every 2 sec. through 20 increasing steps of alternating current. A bar-press reset the shock to its minimal value. The animals performed under conditions of 0, 21.5, and 45.5 hr. water deprivation. When water deprived, animals showed decrewed sensitivity to shock. Frequency of bar-press responses at each of the 20 shock values served as the dependent measure. Findings were discussed in terms of competing techniques for measurement of analgesia.
Geller and Axelrod (1968) have recently evaluated the use of operant techniques in the study of analgesics. They found reflexive measures (such as the tail flick response) to show high inter-experimenter variability and to be sensitive to compounds which are not usually clinically effective in pain reduction. That is, control animals injected with water, propandiol or saline showed more pain threshold elevation than animals under most dose levels of morphine. After rejecting avoidance and escape-avoidance techniques of pain measurement for similar reasons, they recommended the titration method of Weiss and Laties (1958) as the most efficient. The titration method (or fractional-escape as it is sometimes called) produces an increasing intensity of electric footshock to an animal. Each bar-press the animal makes, however, reduces the shock intensity one increment. Thus, animals "hold" the shock intensity at some threshold level. An alternate method is to have the response reset the shock level to zero. Recently, Houser and VanHart ( 1973) have adopted a shuttle-box escape technique for the evaluation of analgesics. Unfortunately, earlier work has shown pain thresholds derived from this technique to be sensitive to such clinically unresponsive manipulations as water deprivation (Misanin & Campbell, 1969). Reflexive measures of pain (vigorous movements, vocalizations or excessive urination) also appear to be sensitive to negative water loads (Griffiths, 1962). The present study was undertaken to examine the sensitivity of the titration technique to manipulations of water balance.
METHOD Two experimentally naive, male, Sprague-Dawley rats were used as Ss. They were 170 days old at the beginning of the study. Testing occurred in a Lehigh Valley small animal test cage which was inside a sound-insulated, environmental cubicle. White noise was continuously present.
488
W. E. FLYNN, ET AL.
The electrical stimulus was produced by a Foringer constant current shock generator with scrambler. During a portion of the procedure used for scaling, the output of the stimulator varied from 0.27 to 0.91 mA. For the remainder of the experiment, the output range was from 0.55 to 1.88 mA. During both settings, the stimulus was varied in 20 discrete and nearly equal steps. Pulses delivered to a stepping switch raised the intensity of the shock by one step every 2 sec. Whenever the lever in the experimental chamber was depressed, the output of the stimulator was reset to its minimal value. Output from the bar was fed through a pulseformer, so that the bar had to be released before a new response could be made. Counters connected to the stepping device recorded the total number of responses made at each step during 2-hr. sessions. During preuaining, Ss were housed in individual cages in a room illuminated on a 12-hr. dark, 12-hr. light cycle. The temperature and humidity were approximately constant. Throughout the experiment, Ss were allowed ad libitam access to Purina Laboratory Chow Checkers. Ss were allowed 30-min. daily access to water. More than 2 mo. of pretraining with 30-min. access to water stabilized daily body weights and water inmke before the experiment began. Ss were shaped to bar-press by presenting a .5-mA shock which was eliminated for 45 sec. by a response. After this response was shaped into a bar-press with a latency of less than 45 sec. from shock onset, 2 hr. daily titration sessions were initiated. Sessions were conducted 7 days per week, 2 hr. per day. The purpose of the first 2 stages of the experiment was to examine the measure of shock sensitivity. Both stages were conducted with Ss 0-ht. water deprived, that is, their 30-min. access to water occurred immediately prior to the 2-hr. experimental session. Ss were tested at a low shock range (0.27 to 0.91 mA) for 16 days. During this period, their performance stabilized. For the second stage of the scaling process, and for the remainder of the experiment, the output of the shock generator was set on the higher ranges (0.55 to 1.88 mA). Measures were taken at this stage for 22 days, during which time performance again stabilized. This latter stage also served as the 0-hr. deprived stage. The procedure for the 21.5-hr. deprived stage was identical to the previous 2 stages except for the watering schedule. During this stage, Ss received water immediately after each day's experimental session. In this way, Ss were 21.5-hr. deprived when the next session started. Data were collected for 11 days under this condition. For the following 24 days, Ss received water every other day immediately after an experimental session. A measure of the aversiveness of the shock was taken, however, every day. This gave 12 additional measures of aversiveness at 2 1.5-hr. deprivation, interspersed with 12 measures of aversiveness at 45.5-hr. deprivation. The final condition was a reinstatement of an earlier stage. For these 10
WATER BALANCE AND TITRATED PAIN THRESHOLDS
489
days, the animals' performance was measured immediately after having access to water.
RESULTS For each S the number of responses made at each of the 20 steps of increasing shock intensity was recorded for every experimental session. The measure of shock sensitivity used was a ratio of the number of responses made in the first 10 steps divided by the number of responses made in the last 10 steps. Thus, higher ratios indicate greater sensitivity to shock. Sensitivity ratios for the last 5 days in each treatment condition are shown in Fig. 1 for both Ss. Inspection of the figure suggests two conclusions. First, the sensitivity ratio appears to be responsive to increases in shock intensity. During the scaling procedure (Panels A and B) , this value increased when the electrical stimulus was changed from the low to the high shock range. Secondly, Ss' sensitivity to shock decreased when they were water deprived as indicated by B and E vs C and D. An analysis of variance performed on these dam confirmed a significant over-all treatment effect for each S (+ < .05). A 5 .
4 .
I
-
8
P
D
C
E
a1.5h..
N
Y : , 1
A
45.5 hr.
I . LO SHOCK H I SMOCK
0
0
H I SHOCK
11.5
HI
SHOCK
21.5 m d 45.5
H I SHOCX
0
HOURS OF WATER DEPRIVATION
FIG. 1. Percentage of Ss' responses that terminated low rather than high shock intensities. Panels B through D show effect of 0-, 21.5- and 45.5-hr. water deprivation. During the 45.5-hr. deprivation period, shack sensitivity was also determined on preceding days (Panel D, 21.5 hr.). The first panel ( A ) shows, for comparison, a lower range of shock output from the source.
W.
490
E. FLYNN, ET AL.
A Newman-Keuls contrast procedure was used to examine individual differences between the various experimental conditions. The scaling procedure which compared low vs high ranges of shock (Panel A vs B ) showed significant differences in the expected direction. That is, Ss responded more often on lower shock steps to the more intensive shock range. For both Ss, the difference between 0-hr. deprivation (Panel B) and both 21.5-hr. (Panel C ) and 45.5-hr. deprivation (Panel D ) proved to be significant (p .05). When Ss were deprived of water for 45.5 hr., however, sensitivity to shock was not significantly different (p > .05) from 21.5-hr. deprivation (Panel C vs D ) . Within the water-every-other-day period (Panel D) sensitivity was significantly lower during the 45.5-hr. deprived segment than during the 21.5-hr. deprived segment ( p < .05). A significant difference ( p < .05) was observed for both Ss during the 21.5-hr. deprived condition (Panel C ) compared to 21.5-hr. deprivation during the alternating 21.5- and 45.5-hr. deprived condition (Panel D) . N o significant differences ( p > .O5) were found between the second 0-hr. deprived condition and the return to that condition (Panel B vs E ) . Thus, manipulations of the independent variable of water deprivation did not appear to affect Ss' original reaction to shock at 0-hr. deprivation. Any fluctuations from the baseline value shown in Panels B and E which might be observed in the remaining panels should, therefore, represent some manipulation of water balance and not a simple effect of trials or learning. It should be noted thac water balance in the present study modified pain thresholds, but in some non-monotonic manner. For example, the 45.5-hr. deprivation condition failed to differ significantly from 21.5 hr. in Panel D. Moreover, during 21.5-hr. water deprivation, sensitivity was lower when water was offered daily than when water was given every 2 days. These discrepancies should be viewed in light of compensatory drinking behaviors observed during the 48-hr. deprivation period (see Fig. 2 ) . That is, animals apparently learned to drink and store more water under 45.5-hr. deprivation. Thus, water deficits in the 45.5-hr. condition may have been no different from the 21.5-hr. condition. Bolles (1967) has suggested thac rats will "learn to drink" much as they have been shown to learn to eat when deprived of food (Hebb, 1949; Reid & Finger, 1955). This means that the animal would learn to compensate for any existing deprivation state by overdrinking during those periods when water was made available. Fig. 2 gives the milliliters of water consumed and the body weights during the last 5 days of each condition for both Ss. An analysis of variance performed on these data indicated a significant treatment effect over conditions for both Ss ( p < .05). A Newman-Keuls contrast procedure showed thac Ss consumed significantly more water ( p .O5) when allowed access to water only on every other day (Panel D ) as opposed to every day (Panels A, B, C, and
