- Email: [email protected]
Self-administration of ethanol by genetically heterogeneous mice (RU:NCS): Relationship to sensitivity and tolerance
Drug and Alcohol Dependence, 12 (1983) Elsevier Scientific Publishers Ireland Ltd.
333
333-338
SELF-ADMINISTRATION OF ETHANOL BY GENETICALLY HETEROGENEOUS MICE (RU: NCS) : RELATIONSHIP TO SENSITIVITY AND TOLERANCE
WILL J. MILLARD* Laboratory of Behavioral Medicine and Pharmacology, Mount Holyoke College, South Hadley, MA 01075 and the Rockefeller University Hospital, New York, NY 10021 (U.S.A.) (Received August 26th, 1983)
SUMMARY
The interrelationships among sensitivity, tolerance and the self-administration of ethanol were studied in a non-inbred strain of mice. Blood ethanol concentration at the time of righting-loss following a hypnotic dose of ethanol (3.5 g/kg) was measured before and after 8 weeks exposure to a choice of water and ethanol water drinking solutions. The inter-test difference between blood ethanol concentration at loss of righting indicated that self-administration resulted in chronic tolerance. A correlational analysis indicated chronic tolerance, but not sensitivity, was positively correlated with ethanol intake and preference. Key words: Blood ethanol concentration Sleep time - Righting reflex - Tolerance
- Ethanol
- Mice - Sensitivity
-
Tolerance is of interest in the study of addictions because the altered action or disposition of a drug may increase self-administration [l] and the resulting physiopathology [ 231. With respect to ethanol, variation of initial sensitivity and tolerance is associated with preference for ethanol in selfadministration procedures [4,5], and this association has been related to differences in the pharmacokinetics of ethanol [6--81 and sensitivity to ethanol [8-111. Chronic tolerance is a result of repeated drug exposure and has been considered a necessary aspect for the development of a non-human analogue of ethanol addiction [ 121. Unlike initial sensitivity and acute tolerance, the relationship between chronic tolerance and ethanol preference remains unestablished. The present experiment determined whether chronic tolerance *Reprint requests to: W.J. Millard, The Rockefeller University Hospital, 1230 York Avenue, New York, NY 10021, U.S.A. 0376-8716/83/$03.00 o 1983 Elsevier Scientific Publishers Ireland Ltd Printed and Published in Ireland
334
accrues as a result of ethanol selection and, by use of a correlational analysis, further examined the degree of association between m.easures of initial sensitivity, tolerance, and ethanol intake and preference. MATERIALS
Subjects
AND METHODS
and apparatus
Thirty-two male mice of the RU : NCS(S) strain were obtained from the Laboratory Animal Research Center of The Rockefeller University. This albino, non-inbred, strain is derived from the original ‘Swiss’ group imported to the United States [ 131. The mice, approx. 9 weeks of age, were received in two shipments of 16 animals for the two replications of the procedure. Each mouse was placed in a polystyrene cage (Laboratory Products) with hardwood litter, and given access to food (#5102, Ralston Purina) and acidified water. The cages were placed in a temperature-controlled vivarium (22 f 2”C), lighted according to a 12/12 h light/dark schedule (on 05: 00 h EST). The water (W), 10% (ElO), and 20% (E20) ethanol-water solutions (v/v, absolute ethanol, U.S.I.) were presented in 100 ml graduated glass bottles fitted with rubber stoppers and stainless steel tubes (Wahmann). Drinking was measured by solid-state lick detectors, and the data were recorded by a microprocessor-baaed computer system [ 14-16 ] . The solutions, bottles, and hardwood shavings were replaced at 7day intervals between 10:00 and 11:OO h EST. Body weight (kO.1 g) and fluid volumes (+0.5 ml) were recorded during this period. Procedure
Animals were initially tested for sensitivity to ethanol by measuring righting-loss following an injection of ethanol [ 61. Sensitivity was defined as the blood ethanol concentration at loss of righting (BECK). The mice were randomly assigned to two conditions (n = 8 per condition/replication) and were given access to either water and the two ethanol-water solutions for 8 weeks (EtOH condition), or three bottles of water for 8 weeks (No-EtOH condition). The ethanol solutions were removed from the cages of the EtOH mice at the end of this period and the second test of righting was administered 24 h later. Inter-test comparisons of the BECL (i.e., ABECL) and duration of righting-loss (i.e ., A duration), served as measures of chronic tolerance [ 3] . The righting test began between 18: 00 and 22 : 00 h with an injection of ethanol (3.5 g/kg body wt. i.p. 20% w/v in distilled water). The mouse was placed on its back in a V-shaped acrylic trough. Failure to right indicated righting-loss, and two subsequent rightings within 20 s defined regaining of righting. The times from injection to loss of righting and regaining of righting were recorded. Blood samples were obtained from the retro-orbital plexus at the loss of righting [ 171. A modification of the gas chromatographic technique described by Freund [ 181 was used for the determination of BEC
335
(7620 A chromatograph, 3390A integrator, Hewlett-Packard; Chromosorb 101 column packing, Supelco). Three determinations were made for each plasma sample. The analysis of variance was used with condition (EtOH vs. No-EtOH) and replication (1 vs. 2) as between-subjects factors, and, for the measures of BECL and duration of righting-loss test (1 vs. 2) as a within-subjects factor. The Bonferroni t was used for pair-wise comparisons of means [ 191, and the method of partial correlation [ 201 was applied to study the association among sensitivity, tolerance, preference and intake of ethanol, with the linear effect of body wt. removed. RESULTS
Ethanol self-administration and tolerance The BEC and duration data from the two tests of righting are presented in Table I with, for the EtOH group, measures of ethanol intake and preference for different periods (Day 1, Week 1, Week 8). Analysis of the BEC, TABLE I MEAN (S.E.M.) MEASURES OF RIGHTING-TEST PERFORMANCE AND ETHANOL PREFERENCE (g EtOH/kg water - day) AND INTAKE (g EtOH/kg body wt. - day) BEC is expressed as mg/dl; durations are in min. Experimental condition
Measure
EtOH
Duration (min.) BECL Chronic tolerance A BECL A Duration
No-EtOH
Test 1
Test 2
Test 1
Test 2
18.5 (4.3) 389 (9)
19.0 (2.3) 477 (22)
14.4 (4.2) 386 (16)
17.1 (2.8) 412 (12)a
(Test I -Test 2)
EtOH preference Day 1 Week 1 Week 8
88 (28) -4.3 (1.9)
26 -5.1
(9)b (3.4)
30.2 (11.4) 35.7 (5.9) 45.0 (6.8)
-
EtOH intake Day 1 Week 1 Week 8
3.2 (0.7) 4.7 (0.7) 5.1 (0.7)
-
abody
7.2 (0.3)
6.3 (0.2)
wt., Week 8-Week 1
aMain effect: EtOH vs. No-EtOH, P < 0.02; Test 1 vs. Test 2, P < 0.001. bMain effect: EtOH vs. No-EtOH. P < 0.01.
-
336
data indicated significant main effects for condition [F,, ssj = 5.67, P < 0.021 and test [Fo.28j = 37.38, P < O.OOl],and a significant interaction of the main effects [F(, ss) = 11.04, P < 0.0051. The effect of replication in all analyses was not &nificant. Pair-wise comparisons showed the mean BECr, for the EtOH and No-EtOH groups for Test 1 were not significantly different and that the BECL in the second test increased for both groups [EtOH, ttzsj = 6.57, P < 0.01; No-EtOH, tczsj = 1.97, P < 0.051. In Test 2 the BECL for the EtOH group was significantly higher than the No-EtOH group [t(28j = 4.85, P < 0.011, indicating the development of chronic tolerance as a result of ethanol intake. Duration of righting-loss was unaffected by either condition [F (1,28)= 0.04, NSI, or test [FcI,28j= 1.01, NS] , and the interaction of main effects was not significant [Ft1,2sj = 0.97, NS]. Mean A body wt. was not reliably altered by the treatment [F,,,as, = 0.73, NS] . E tOH mice:
correlational
analysis
Variables predicting high preference for ethanol were sought in the partial correlation analysis of the measures of sensitivity, tolerance, and ethanol self-administration, the results of which are presented in Table II. Initial sensitivity was not significantly associated with the asymptotic measures of
TABLE
II
CORRELATION MATRIX FOR MEASURES OF SENSITIVITY, TOLERANCE AND ETHANOL INTAKE (g EtOH/kg water * day) AND PREFERENCE (g EtOH/kg water - day) Values
are partial correlation
Measure
1
1. Sensitivity (BEQ Test 1)
Chronic tolerance (Test 1 -Test 2) 2. aBECL 3.
A
Duration
EtOH intake 4. Day 1 5. Week 1 6. Week 8
coefficients, 2
3
-0.01
-
with the linear effect
of body
4
5
6
-0.07
-
wt. removed. 7
8
9
0.31 0.36
0.26
-
-
0.09 0.22
-0.17 -0.10 0.21
0.13 0.37 0.85a
-0.18 0.31 -0.04
0.06 0.13
-0.18 -0.10 0.18
0.12 0.30 0.84’
-0.29 0.29 -0.22
0.91* 0.03 0.10
EtOH preference 7.Dayl 8. Week 1 9. Week 8 ar > 0;P
< 0.01.
0.06 0.98a 0.11
0.13 -0.10 0.84*
331
intake. Chronic tolerance, however, as measured by ABEQ,, but not A duration, was positively correlated with intake (r = 0.85) and preference (F‘= 0.84) dur ing the last week of self-administration. DISCUSSION
The results of the experiment indicate the self-administration of ethanol resulted in chronic tolerance as measured by higher levels of ethanol in blood at loss of righting. Prior research has established the relationship between ethanol intake and the development of tolerance [ 211. Unlike previous methods in which ethanol administration was forced, the mice in the present experiment acquired chronic tolerance to the hypnotic effect of ethanol as a result of self-administration in a choice procedure, and without concomitant food or water deprivation. The correlational analysis showed the degree of chronic tolerance was positively correlated with ethanol intake at the end of the &week period of self-administration. Under the present experimental conditions the intake of approx. 5 g EtOH/kg body wt. * day resulted in a measurable degree of tolerance. This amount for rodents, however, does not result in enduring intoxication, withdrawal symptoms or pathology [12,16]. Thus, the chronic tolerance associated with the observed levels of ethanol intake suggests tolerance is not a sufficient antecedent or correlate of ethanol addition. ACKNOWLEDGEMENTS
This investigation was supported by grants to The Rockefeller University from the U.S. Public Health Service (F32 AA05169), John L. and Helen Kellogg Foundation, the Florina Lasker Charitable Trust, and by a grant to Mount Holyoke College from the Merck Foundation. The gracious support of Vincent P. Dole, M.D., Head, Biology of Addictive Diseases of The Rockefeller University, is gratefully acknowledged. The modified gas chromatographic technique was developed by Michael S. Rappaport, Ph.D. The technical assistance of Adrienne J. Chin and the editorial advice of R. Thomas Gentry, Ph.D., Ann Ho, Ph.D. and Michael S. Rappaport, Ph.D., is appreciated. REFERENCES 1 J.H. Jaffe, in: Gilman, L.S. Goodman and A. Gilman A (Eds.), Goodman and Gilman’s: The Pharmacological Basis of Therapeutics, Macmillan, New York, 1981, pp. 535584. 2 H. Cappell and A.E. LeBlanc, Drug Alcohol Depend., 4 (1979) 15. 3 B. Tabakoff, in: K. Eriksson, J.D. Sinclair and K. Kiianmaa (Eds.), Animal Models in Alcohol Research, Academic, New York, 1980, pp. 271-292. 4 V.G. Erwin, G.E. McClearn and A.R. Kuse, Pharmacol. Biochem. Behav., 13 (1980) 297. 5 G.E. McClearn and D.A. Rodgers, J. Comp. Physiol. Psychol., 54 (1961) 116.
338 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
J.K. Belknap et al., Physiol. Behav., 9 (1972) 453. JR. Sheppard, P. Albersheim and G. McClearn, J. Biol. Chem., 245 (1970) 2876. C.W. Schneider et al., J. Comp. Physiol. Psychol., 82 (1973) 466. R. Damjanovich and J. MacInnes, Life Sci., 13 (1973) 55. R. Kakihana, D.R. Brown and G.E. McClearn, Science, 154 (1966) 1574. P. Nikander and L. Pekkanen, Psychopharmacology, 51 (1977) 219. D. Lester and E.X. Freed, Pharmacol. Biochem. Behav., 1 (1973) 103. C.J. Lynch, Lab. Anim. Care., 19 (1969) 214. V.P. Dole, A. Ho and R.T. Gentry, Physiol. Behav., 30 (1983) 971. R.T. Gentry, M.S. Rappaport and V.P. Dole, Physiol. Behav., 31 (1983) in press. W.J. Millard and V.P. Dole, Pharmacol. Biochem. Behav., 18 (1983) 281. V. Riley, Proc. Sot. Exp. Biol. Med., 104 (1960) 751. G. Freund, Anal. Chem., 39 (1967) 545. J.L. Myers, Fundamentals of Experimental Design, Allyn and Bacon, Boston, 1979. J. Cohen and P. Cohen, Applied Multiple Regression/Correlation Analysis for the Behavioral Sciences, Erlbaum, Hillsdale, New Jersey, 1975. 21 H. Kalant, A.E. LeBlanc and R.J. Gibbins, Pharmacol. Rev., 23 (1971) 135.
333
333-338
SELF-ADMINISTRATION OF ETHANOL BY GENETICALLY HETEROGENEOUS MICE (RU: NCS) : RELATIONSHIP TO SENSITIVITY AND TOLERANCE
WILL J. MILLARD* Laboratory of Behavioral Medicine and Pharmacology, Mount Holyoke College, South Hadley, MA 01075 and the Rockefeller University Hospital, New York, NY 10021 (U.S.A.) (Received August 26th, 1983)
SUMMARY
The interrelationships among sensitivity, tolerance and the self-administration of ethanol were studied in a non-inbred strain of mice. Blood ethanol concentration at the time of righting-loss following a hypnotic dose of ethanol (3.5 g/kg) was measured before and after 8 weeks exposure to a choice of water and ethanol water drinking solutions. The inter-test difference between blood ethanol concentration at loss of righting indicated that self-administration resulted in chronic tolerance. A correlational analysis indicated chronic tolerance, but not sensitivity, was positively correlated with ethanol intake and preference. Key words: Blood ethanol concentration Sleep time - Righting reflex - Tolerance
- Ethanol
- Mice - Sensitivity
-
Tolerance is of interest in the study of addictions because the altered action or disposition of a drug may increase self-administration [l] and the resulting physiopathology [ 231. With respect to ethanol, variation of initial sensitivity and tolerance is associated with preference for ethanol in selfadministration procedures [4,5], and this association has been related to differences in the pharmacokinetics of ethanol [6--81 and sensitivity to ethanol [8-111. Chronic tolerance is a result of repeated drug exposure and has been considered a necessary aspect for the development of a non-human analogue of ethanol addiction [ 121. Unlike initial sensitivity and acute tolerance, the relationship between chronic tolerance and ethanol preference remains unestablished. The present experiment determined whether chronic tolerance *Reprint requests to: W.J. Millard, The Rockefeller University Hospital, 1230 York Avenue, New York, NY 10021, U.S.A. 0376-8716/83/$03.00 o 1983 Elsevier Scientific Publishers Ireland Ltd Printed and Published in Ireland
334
accrues as a result of ethanol selection and, by use of a correlational analysis, further examined the degree of association between m.easures of initial sensitivity, tolerance, and ethanol intake and preference. MATERIALS
Subjects
AND METHODS
and apparatus
Thirty-two male mice of the RU : NCS(S) strain were obtained from the Laboratory Animal Research Center of The Rockefeller University. This albino, non-inbred, strain is derived from the original ‘Swiss’ group imported to the United States [ 131. The mice, approx. 9 weeks of age, were received in two shipments of 16 animals for the two replications of the procedure. Each mouse was placed in a polystyrene cage (Laboratory Products) with hardwood litter, and given access to food (#5102, Ralston Purina) and acidified water. The cages were placed in a temperature-controlled vivarium (22 f 2”C), lighted according to a 12/12 h light/dark schedule (on 05: 00 h EST). The water (W), 10% (ElO), and 20% (E20) ethanol-water solutions (v/v, absolute ethanol, U.S.I.) were presented in 100 ml graduated glass bottles fitted with rubber stoppers and stainless steel tubes (Wahmann). Drinking was measured by solid-state lick detectors, and the data were recorded by a microprocessor-baaed computer system [ 14-16 ] . The solutions, bottles, and hardwood shavings were replaced at 7day intervals between 10:00 and 11:OO h EST. Body weight (kO.1 g) and fluid volumes (+0.5 ml) were recorded during this period. Procedure
Animals were initially tested for sensitivity to ethanol by measuring righting-loss following an injection of ethanol [ 61. Sensitivity was defined as the blood ethanol concentration at loss of righting (BECK). The mice were randomly assigned to two conditions (n = 8 per condition/replication) and were given access to either water and the two ethanol-water solutions for 8 weeks (EtOH condition), or three bottles of water for 8 weeks (No-EtOH condition). The ethanol solutions were removed from the cages of the EtOH mice at the end of this period and the second test of righting was administered 24 h later. Inter-test comparisons of the BECL (i.e., ABECL) and duration of righting-loss (i.e ., A duration), served as measures of chronic tolerance [ 3] . The righting test began between 18: 00 and 22 : 00 h with an injection of ethanol (3.5 g/kg body wt. i.p. 20% w/v in distilled water). The mouse was placed on its back in a V-shaped acrylic trough. Failure to right indicated righting-loss, and two subsequent rightings within 20 s defined regaining of righting. The times from injection to loss of righting and regaining of righting were recorded. Blood samples were obtained from the retro-orbital plexus at the loss of righting [ 171. A modification of the gas chromatographic technique described by Freund [ 181 was used for the determination of BEC
335
(7620 A chromatograph, 3390A integrator, Hewlett-Packard; Chromosorb 101 column packing, Supelco). Three determinations were made for each plasma sample. The analysis of variance was used with condition (EtOH vs. No-EtOH) and replication (1 vs. 2) as between-subjects factors, and, for the measures of BECL and duration of righting-loss test (1 vs. 2) as a within-subjects factor. The Bonferroni t was used for pair-wise comparisons of means [ 191, and the method of partial correlation [ 201 was applied to study the association among sensitivity, tolerance, preference and intake of ethanol, with the linear effect of body wt. removed. RESULTS
Ethanol self-administration and tolerance The BEC and duration data from the two tests of righting are presented in Table I with, for the EtOH group, measures of ethanol intake and preference for different periods (Day 1, Week 1, Week 8). Analysis of the BEC, TABLE I MEAN (S.E.M.) MEASURES OF RIGHTING-TEST PERFORMANCE AND ETHANOL PREFERENCE (g EtOH/kg water - day) AND INTAKE (g EtOH/kg body wt. - day) BEC is expressed as mg/dl; durations are in min. Experimental condition
Measure
EtOH
Duration (min.) BECL Chronic tolerance A BECL A Duration
No-EtOH
Test 1
Test 2
Test 1
Test 2
18.5 (4.3) 389 (9)
19.0 (2.3) 477 (22)
14.4 (4.2) 386 (16)
17.1 (2.8) 412 (12)a
(Test I -Test 2)
EtOH preference Day 1 Week 1 Week 8
88 (28) -4.3 (1.9)
26 -5.1
(9)b (3.4)
30.2 (11.4) 35.7 (5.9) 45.0 (6.8)
-
EtOH intake Day 1 Week 1 Week 8
3.2 (0.7) 4.7 (0.7) 5.1 (0.7)
-
abody
7.2 (0.3)
6.3 (0.2)
wt., Week 8-Week 1
aMain effect: EtOH vs. No-EtOH, P < 0.02; Test 1 vs. Test 2, P < 0.001. bMain effect: EtOH vs. No-EtOH. P < 0.01.
-
336
data indicated significant main effects for condition [F,, ssj = 5.67, P < 0.021 and test [Fo.28j = 37.38, P < O.OOl],and a significant interaction of the main effects [F(, ss) = 11.04, P < 0.0051. The effect of replication in all analyses was not &nificant. Pair-wise comparisons showed the mean BECr, for the EtOH and No-EtOH groups for Test 1 were not significantly different and that the BECL in the second test increased for both groups [EtOH, ttzsj = 6.57, P < 0.01; No-EtOH, tczsj = 1.97, P < 0.051. In Test 2 the BECL for the EtOH group was significantly higher than the No-EtOH group [t(28j = 4.85, P < 0.011, indicating the development of chronic tolerance as a result of ethanol intake. Duration of righting-loss was unaffected by either condition [F (1,28)= 0.04, NSI, or test [FcI,28j= 1.01, NS] , and the interaction of main effects was not significant [Ft1,2sj = 0.97, NS]. Mean A body wt. was not reliably altered by the treatment [F,,,as, = 0.73, NS] . E tOH mice:
correlational
analysis
Variables predicting high preference for ethanol were sought in the partial correlation analysis of the measures of sensitivity, tolerance, and ethanol self-administration, the results of which are presented in Table II. Initial sensitivity was not significantly associated with the asymptotic measures of
TABLE
II
CORRELATION MATRIX FOR MEASURES OF SENSITIVITY, TOLERANCE AND ETHANOL INTAKE (g EtOH/kg water * day) AND PREFERENCE (g EtOH/kg water - day) Values
are partial correlation
Measure
1
1. Sensitivity (BEQ Test 1)
Chronic tolerance (Test 1 -Test 2) 2. aBECL 3.
A
Duration
EtOH intake 4. Day 1 5. Week 1 6. Week 8
coefficients, 2
3
-0.01
-
with the linear effect
of body
4
5
6
-0.07
-
wt. removed. 7
8
9
0.31 0.36
0.26
-
-
0.09 0.22
-0.17 -0.10 0.21
0.13 0.37 0.85a
-0.18 0.31 -0.04
0.06 0.13
-0.18 -0.10 0.18
0.12 0.30 0.84’
-0.29 0.29 -0.22
0.91* 0.03 0.10
EtOH preference 7.Dayl 8. Week 1 9. Week 8 ar > 0;P
< 0.01.
0.06 0.98a 0.11
0.13 -0.10 0.84*
331
intake. Chronic tolerance, however, as measured by ABEQ,, but not A duration, was positively correlated with intake (r = 0.85) and preference (F‘= 0.84) dur ing the last week of self-administration. DISCUSSION
The results of the experiment indicate the self-administration of ethanol resulted in chronic tolerance as measured by higher levels of ethanol in blood at loss of righting. Prior research has established the relationship between ethanol intake and the development of tolerance [ 211. Unlike previous methods in which ethanol administration was forced, the mice in the present experiment acquired chronic tolerance to the hypnotic effect of ethanol as a result of self-administration in a choice procedure, and without concomitant food or water deprivation. The correlational analysis showed the degree of chronic tolerance was positively correlated with ethanol intake at the end of the &week period of self-administration. Under the present experimental conditions the intake of approx. 5 g EtOH/kg body wt. * day resulted in a measurable degree of tolerance. This amount for rodents, however, does not result in enduring intoxication, withdrawal symptoms or pathology [12,16]. Thus, the chronic tolerance associated with the observed levels of ethanol intake suggests tolerance is not a sufficient antecedent or correlate of ethanol addition. ACKNOWLEDGEMENTS
This investigation was supported by grants to The Rockefeller University from the U.S. Public Health Service (F32 AA05169), John L. and Helen Kellogg Foundation, the Florina Lasker Charitable Trust, and by a grant to Mount Holyoke College from the Merck Foundation. The gracious support of Vincent P. Dole, M.D., Head, Biology of Addictive Diseases of The Rockefeller University, is gratefully acknowledged. The modified gas chromatographic technique was developed by Michael S. Rappaport, Ph.D. The technical assistance of Adrienne J. Chin and the editorial advice of R. Thomas Gentry, Ph.D., Ann Ho, Ph.D. and Michael S. Rappaport, Ph.D., is appreciated. REFERENCES 1 J.H. Jaffe, in: Gilman, L.S. Goodman and A. Gilman A (Eds.), Goodman and Gilman’s: The Pharmacological Basis of Therapeutics, Macmillan, New York, 1981, pp. 535584. 2 H. Cappell and A.E. LeBlanc, Drug Alcohol Depend., 4 (1979) 15. 3 B. Tabakoff, in: K. Eriksson, J.D. Sinclair and K. Kiianmaa (Eds.), Animal Models in Alcohol Research, Academic, New York, 1980, pp. 271-292. 4 V.G. Erwin, G.E. McClearn and A.R. Kuse, Pharmacol. Biochem. Behav., 13 (1980) 297. 5 G.E. McClearn and D.A. Rodgers, J. Comp. Physiol. Psychol., 54 (1961) 116.
338 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
J.K. Belknap et al., Physiol. Behav., 9 (1972) 453. JR. Sheppard, P. Albersheim and G. McClearn, J. Biol. Chem., 245 (1970) 2876. C.W. Schneider et al., J. Comp. Physiol. Psychol., 82 (1973) 466. R. Damjanovich and J. MacInnes, Life Sci., 13 (1973) 55. R. Kakihana, D.R. Brown and G.E. McClearn, Science, 154 (1966) 1574. P. Nikander and L. Pekkanen, Psychopharmacology, 51 (1977) 219. D. Lester and E.X. Freed, Pharmacol. Biochem. Behav., 1 (1973) 103. C.J. Lynch, Lab. Anim. Care., 19 (1969) 214. V.P. Dole, A. Ho and R.T. Gentry, Physiol. Behav., 30 (1983) 971. R.T. Gentry, M.S. Rappaport and V.P. Dole, Physiol. Behav., 31 (1983) in press. W.J. Millard and V.P. Dole, Pharmacol. Biochem. Behav., 18 (1983) 281. V. Riley, Proc. Sot. Exp. Biol. Med., 104 (1960) 751. G. Freund, Anal. Chem., 39 (1967) 545. J.L. Myers, Fundamentals of Experimental Design, Allyn and Bacon, Boston, 1979. J. Cohen and P. Cohen, Applied Multiple Regression/Correlation Analysis for the Behavioral Sciences, Erlbaum, Hillsdale, New Jersey, 1975. 21 H. Kalant, A.E. LeBlanc and R.J. Gibbins, Pharmacol. Rev., 23 (1971) 135.