- Email: [email protected]
Changes in defensive behaviors following olfactory bulbectomy in male and female rats
Brain Research 903 (2001) 242–246 www.elsevier.com / locate / bres
Short communication
Changes in defensive behaviors following olfactory bulbectomy in male and female rats Howard S. Stock
a,b ,
*, Gregory A. Hand a , Kris Ford a , Marlene A. Wilson a
a
University of South Carolina, School of Medicine, Department of Pharmacology and Physiology, Columbia, SC 29208, USA b WJB Dorn Department of Veterans Affairs Medical Center, 6439 Garners Ferry Road, Columbia, SC 29209 -1439, USA Accepted 20 March 2001
Abstract The present study examined if olfactory bulbectomy (OBX) altered defensive behaviors on the elevated plus-maze and the open-field differently in male and female rats. Similar increases in defensive behaviors in male and female rats were observed in both tests following OBX. No significant correlations were detected between defensive behaviors and activity, supporting the hypothesis that some behavioral changes following OBX may be due to decreased defensive behaviors and not increased activity. 2001 Elsevier Science B.V. All rights reserved. Theme: Neural basis of behavior Topic: Stress Keywords: Sex difference; Defensive behavior; Plus-maze; Open-field; Model of depression
1. Introduction In stressful situations animals will elicit a variety of defensive behaviors, including: freezing, analgesia, burying, escape, and avoidance [1]. These behaviors are enhanced in novel environments and are attenuated in familiar environments. Moreover, the attenuation and enhancement of these behaviors can also be modulated by anxiolytic or anxiogenic compounds, respectively. The elevated plus-maze test, used mainly as a measure of anxiety in rodents, is based on the approach–avoidance aspect of a rat’s defensive response. It has been proposed that the open arms of the plus-maze produce a greater aversive reaction because they have a greater amount of novel stimulation compared to the closed arms (see [4] for references). Therefore, an increase in open arm time is suggestive of a decrease in a rat’s defensive responding. In addition, measuring exploration behavior the open-field test has also been used as a measure of defensive behaviors *Corresponding author. Tel.: 11-803-733-3156; fax: 11-803-7331523. E-mail address: [email protected] (H.S. Stock).
in rodents, where increased line crossings and rearing are suggestive of a decrease in defensive behaviors [14,19]. In the rat, removal of the olfactory bulbs results in a wide variety of behavioral and neurochemical changes including decreased serotonin levels, decreased norepinephrine levels, increased corticosterone levels, and increased gama-aminobutyric acid (GABA) levels [8]. Some of the behavioral changes associated with olfactory bulbectomy (OBX), such as increased open-field activity levels, decrements in passive avoidance tasks, increased time spent in open arms in the elevated plus-maze [8], decreased shock-induced freezing, and decreased time spent in a covered enclosure, are thought to be associated with a decrement in the animal’s defensive responding and not due to an increase in activity levels [11]. The majority of previous research examining OBX-induced changes in defensive behaviors have predominantly used male rats. Recently, we have observed sex and gonadal hormone effects on open-field behavior and sucrose preference tasks following OBX [16]. In addition, sex and gonadectomy effects have been observed in GABA–benzodiazepine (BZ) reactivity [20], a system important in modulating defensive responding [3,9,12,13]. Therefore, the present
0006-8993 / 01 / $ – see front matter 2001 Elsevier Science B.V. All rights reserved. PII: S0006-8993( 01 )02421-0
H.S. Stock et al. / Brain Research 903 (2001) 242 – 246
study was designed to determine if the sex and the gonadal status of the animal interact with OBX to alter defensive responding as measured by changes in elevated plus-maze or open-field related behaviors.
2. Methods
2.1. Subjects Male and female Long–Evans hooded rats (Harlan Sprague–Dawley Inc, Indianapolis, IN, USA) were housed singly with food and water available ad libitum. The animals were kept on a 12–12 h light–dark cycle, lights on at 7:00 am, with behavioral testing occurring in the early part of the light phase (0900–1200 h). Animals were housed in an AAALAC-approved animal colony room and all procedures were approved by the University of South Carolina Animal Care and Use Committee.
2.2. Handling and surgeries Handling and surgery techniques were identical to those described in Ref. [16]. Eight groups were tested: male sham (N56), male OBX (N56), orchidectomized (ORCH) sham (N56), OBX–ORCH (N57), female sham (N56), female OBX (N56), ovariectomized (OVX) sham (N55), and OBX–OVX (N55).
2.3. Apparatus and test procedure To determine the interactions between the sex, gonadal status of the animal, and OBX on behavioral responding in a novel open-field animals were tested 2 weeks following OBX in a open-field apparatus as described in Ref. [7]. The animals were placed into the center of the open-field and allowed to explore freely for 5 min. Behavior was video-taped via a video camera suspended 6 feet above the center of the maze. The video tapes were scored at a later date by an experimenter who was blind to the treatment conditions of the animals. The behaviors of interest were the number of lines crossed and the number of rears made during exposure to the open-field. A line crossing was defined as all four paws crossing over the line. Changes in the defensive behaviors following OBX were also assessed using an elevated plus-maze identical to that described in Stock et al. [16]. Twenty-four hours following open-field testing animals were placed into the center square and allowed to explore the maze for a period of 5 min [10]. Behavior was taped via a video camera suspended 3 feet above the center of the maze. The video tapes were scored at a later date by an experimenter who was blind to the treatment conditions of the animals. The behaviors of interest on the elevated plus-maze were percent open arm time (% open arm time), percent open arm entries (% open arm entries), and center square time.
243
An animal was scored as having entered the arm when all four paws were inside the arm. Percent open arm time was calculated as the total time spent in the open arms divided by the total time spent in all arms of the maze. Percent open arm entries was calculated as the total entries into the open arm divided by the total number of entries into all arms of the maze. Both percent open arm time and percent open arm entries have been reported to be behavioral indices of defensive responding in rats, with increases in time and entries suggesting lower defensive behaviors [4,10]. Center square time has been reported as a reliable measure of activity levels for this type of elevated plusmaze (i.e., ledges on open arms) [2].
2.4. Statistics The influences of sex (male vs. female), gonadectomy (intact vs. gonadectomized), and OBX condition (sham vs. OBX) on plus-maze and open-field related behaviors were compared by a three-way analysis of variance (ANOVA; sex, gonadectomy, and OBX as the between-subject factors). The three-way ANOVA with sex, gonadectomy, and OBX condition revealed no main effects or interactions of gonadectomy on any dependent measures. Consequently, data were collapsed across gonadectomy and analyzed with a two-way analysis of variance (sex and OBX condition). For all statistical analyses, Tukey honestly significant difference tests were used as post-hoc tests. The alpha level for all statistical analysis was 0.05. Pearson product moment correlations were conducted between open-field related behaviors and the behavioral measures recorded on the plus-maze.
3. Results
3.1. Number of lines crossed As seen in Fig. 1A, OBX significantly increased the number of line crossings compared to sham animals. A significant OBX main effect was detected [F(1,43)542.38, P50.001]. A Tukey post-hoc test demonstrated that OBX animals showed significantly greater number of lines crossed than sham controls (P,0.05). Pearson’s product moment correlation reported a significant positive correlation between number of lines crossed and number of rears (r 2 50.80, P50.001), percent open arm time (r 2 50.40, P50.005), and percent open arm entries (r 2 50.32 P5 0.027).
3.2. Number of rears As seen in Fig. 1B, OBX significantly increased the number of rears compared to sham animals. A significant OBX main effect was detected [F(1,43)559.17, P5 0.001]. A Tukey post-hoc test demonstrated that OBX
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H.S. Stock et al. / Brain Research 903 (2001) 242 – 246
Fig. 1. Open-field related behaviors: (A) Each bar represents mean number of lines crossed (6S.E.M.) measured during a 5-min exposure to the open-field apparatus. Animals underwent either olfactory bulbectomy [OBX: dark columns (male N513; female N511)] or sham surgery [sham: open columns (male N512; female N511)]. *OBX animals had significantly greater number of lines crossed than sham controls (P, 0.05). (B) Each bar represents mean number of rears (6S.E.M.) measured during a 5-min exposure to the open-field apparatus. Animals underwent either olfactory bulbectomy [OBX: dark columns (male N513; female N511)] or sham surgery [sham: open columns (male N512; female N511)]. *OBX animals had significantly greater number of rears than sham controls (P,0.05).
animals showed a significantly greater number of rears than sham controls (P,0.05). Pearson’s product moment correlations reported a significant positive correlation between number of rears and percent open arm time (r 2 50.33 P50.022) and percent open arm entries (r 2 5 0.36 P50.010).
3.3. Percent open arm entries As seen in Fig. 2A, OBX significantly increased percent open arm entries compared to sham animals. A significant OBX main effect was detected [F(1,43)510.84, P5 0.002]. A Tukey post-hoc test demonstrated that OBX animals showed significantly greater percent open arm
Fig. 2. Plus-maze related behaviors: (A) Each bar represents mean percent open arm entries (6S.E.M.) measured during a 5-min exposure to the elevated plus-maze. Animals underwent either olfactory bulbectomy [OBX: dark columns (male N513; female N511)] or sham surgery [sham: open columns (male N512; female N511)]. *OBX animals had significantly greater percent open arm entries than sham controls (P, 0.05). (B) Each bar represents mean percent open arm time (6S.E.M.) measured during a 5-min exposure to the elevated plus-maze. Animals underwent either olfactory bulbectomy [OBX: dark columns (male N513; female N511)] or sham surgery [sham: open columns (male N512; female N511)]. *OBX animals had significantly greater percent open arm time than sham controls (P,0.05). (C) Each bar represents mean center square time (6S.E.M.) measured during a 5-min exposure to the elevated plus-maze. Animals underwent either olfactory bulbectomy [OBX: dark columns (male N513; female N511)] or sham surgery [sham: open columns (male N512; female N511)].
entries than sham controls (P,0.05). Pearson’s product moment correlations reported a significant positive correlation between percent open arm entries and percent open arm time (r 2 50.89 P50.001)
H.S. Stock et al. / Brain Research 903 (2001) 242 – 246
3.4. Percent open arm time As seen in Fig. 2B, OBX significantly increased percent open arm time compared to sham animals. A significant OBX main effect was detected [F(1,42)58.50, P50.005]. A Tukey post-hoc test demonstrated that OBX animals showed significantly greater percent open arm time than sham controls (P,0.05).
3.5. Center square time As seen in Fig. 2C, no effect of OBX on center square time was observed.
4. Discussion The present study was conducted to examine the possible interaction between OBX and the gonadal hormone milieu on changes in the rat’s defensive behavior, as measured by the open-field and elevated plus-maze. The results show that OBX increases the number of lines crossed as well as the number of rears in the open-field apparatus compared to sham controls, across all hormone groups. In addition, OBX increased the amount of time and entries on the open arms of the elevated plus maze compared to sham controls, across all hormone groups. The increases in number of lines crossed, rears, open arm time, and open arm entries suggest that OBX decreased defensive responding compared to sham controls. Moreover, this decrease in defensive behaviors in the elevated plus-maze does not appear to be due to changes in activity levels, since alterations in center square time, a more sensitive measure of activity with this particular type of plus-maze, was not observed [2]. Moreover, significant positive correlations were detected between open-field related behaviors (i.e., number of lines crossed and rears) and percent open arm time and entries. These findings suggest a relationship between these two tests in regards to defensive behaviors in rats. In the present study no significant correlations between center square time and number of lines crossed, rears, percent open arm time, and percent open arm entries were detected, suggesting no relationship between these measures of defensive behavior and activity. However, prior research suggests that percent open arm entries may be a more valid measure of the approach– avoidance component of the elevated plus-maze than percent open arm time, since it does not appear to be affected by changes in activity [4]. Overall, these results further support the hypothesis that OBX results in a decrease in the rat’s defensive behavior during exposure to an novel environment [11]. In addition, our findings suggest that modulation of gonadal hormone levels via adult gonadectomy does not alter defensive responding
245
following OBX as measured by the open-field and elevated plus-maze. Previous research has reported that control female rats exhibit greater levels of percent open arm time and entries than control male rats [7,21], although this is not a universal finding [5,15]. These discrepancies in outcomes may be due to a variety of procedural differences between these studies; including structural differences between the testing apparatus and strain of rat tested. Our previous research demonstrated that during exposure to an open-field only OBX-gonadectomized male and female rats exhibited a significant change from baseline 2 weeks following OBX [16]. However, in the present study all OBX animals exhibited an increase in open-field related behaviors. Differences in open-field testing procedures may provide a possible explanation for the discrepancy between these two studies. In the present study, the animals were tested in a novel open-field while in our previous study animals were habituated to the open-field before undergoing OBX surgery (i.e., baseline testing). Therefore, habituation and the gonadal status of the animal may have interacted to alter the increase in open-field related behaviors that have previously been observed following OBX [8]. This hypothesis is supported by the finding that gonadally-intact males that were habituated to an open-field apparatus failed to show an increase in open-field activity levels when tested following OBX surgery [17]. Central administration of a variety of compounds (i.e., benzodiazepines) that augment GABAergic functioning have been reported to alter a multitude of defensive behaviors such as freezing, exploration of a novel environment, food intake in a novel environment, and open arm time and entries [12,13]. In addition, prior research has demonstrated that OBX results in an increase in GABAergic functioning in a variety of brain areas (limbic system, cortex) [6,8,18]. Therefore, OBX may result in an increase in GABAergic activity, which in turn may result in a decrease in the defensive responding that has been observed in the present study and in previous research [8,11]. The results of this study replicate prior work that has demonstrated that OBX decreased defensive responding, without altering activity levels in male rats [8,11], and extends this finding to female, gonadectomized male, and ovariectomized female rats. A likely mediator of the decreased defensive responding following OBX may be increased GABAergic activity, however, further investigation will be needed to validate this hypothesis.
Acknowledgements The authors thank Cynthia Gobbel and Stefany Primeaux for helpful comments on the manuscript. This work was supported by a grant from The National Alliance for the Research on Schizophrenia and Depression.
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References [1] R. Blanchard, D. Blanchard, Anti-predator defense as models of animal fear and anxiety, in: P. Brain, P. Stefano (Eds.), Fear and Defense, Harwood Academic Publishers, London, 1990, pp. 89– 108. [2] C. Fernandes, S.E. File, The influence of open arm ledges and maze experience in the elevated plus maze, Pharmacol. Biochem. Behav. 54 (1996) 31–40. [3] F. Graeff, Neuroanatomy and neurotransmitter regulation of defensive behaviors and related emotions in mammals, Braz. J. Med. Biol. Res. 27 (1994) 811–829. [4] S.L. Handley, J.W. McBlane, An assessment of the elevated maze for studying anxiety and anxiety modulating drugs, J. Pharm. Toxic Methods 29 (1993) 129–138. [5] R.P. W Heinsbroek, F. Van Haaren, N.E. Van De Poll, H.L. Steenbergen, Sex differences in the behavioral consequences of inescapable footshocks depend on time since shock, Physiol. Behav. 49 (1991) 1257–1263. [6] J.D. Hirsch, Mini review the neurochemical sequella of olfactory bulbectomy, Life Sci. 26 (1980) 1551–1559. [7] J. Imhof, Z. Coelho, M. Schmitt, G. Morato, A. Carobrez, Influence of gender and age on performance of rats in the elevated plus maze apparatus, Behav. Brain Res. 56 (1993) 177–180. [8] J.P. Kelly, A.S. Wrynn, B.E. Leonard, The olfactory bulbectomized rat as a model of depression: an update, Pharmacol. Ther. 74 (1997) 299–316. [9] J. Pandossio, M. Brandao, Defensive reactions are counteracted by midazolam and muscimol and elicited by activation of glutamate receptors in the inferior colliculus of rats, Psychopharmacology 142 (1999) 360–368. [10] S. Pellow, P. Chopin, S.E. File, M. Briley, Validation of open:closed
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arm entries in an elevated plus-maze as a measure of anxiety in the rat, J. Neurosci. Methods 14 (1985) 149–167. S. Primeaux, P.V. Holmes, Role of aversively motivated behavior in the olfactory bulbectomy syndrome, Physiol. Behav. 67 (1999) 41–47. A. Ramos, O. Berton, P. Mormede, F. Chaouloff, A multiple-test study of anxiety-related behaviors in six inbred rat strains, Behav. Brain Res. 85 (1997) 57–69. A. Rex, D. Stephens, H. Fink, Anxiolytic action of diazepam and abecarnil in a modified open field test, Pharmacol. Biochem. Behav. 53 (1996) 1005–1011. J.R. Royce, On the construct validity of the open-field measures, Psychol. Bull. 84 (1977) 1098–1106. H.S. Stock, C. Foradori, K. Ford, M.A. Wilson, A lack of tolerance to the anxiolytic effects of diazepam on the plus-maze: comparison of male and female rats, Psychopharmacology 147 (2000) 362–370. H.S. Stock, K. Ford, M.A. Wilson, The effects of gender and gonadal hormones in the olfactory bulbectomy animal model of depression, Pharmacol. Biochem. Behav. 67 (1999) 183–191. P.B. Tiffany, S. Mollenauer, R. Plotnik, M. White, Olfactory bulbectomy: emotional behavior and defense responses in the rat, Physiol. Behav. 22 (1979) 311–317. H. van Riezen, B. E Leonard, Effects of psychotropic drugs on the behavior and neurochemistry of olfactory bulbectomized rats, Pharmacol. Ther. 47 (1990) 21–34. R.N. Walsh, R.A. Cummins, The open-field test: a critical review, Psychol. Bull. 83 (1976) 482–504. M.A. Wilson, GABA physiology: modulation by benzodiazepines and hormones, Crit. Rev. Neurobiol. 10 (1996) 1–37. B. Zimmerberg, M.J. Farley, Sex differences in anxiety behavior in rats: role of gonadal hormones, Physiol. Behav. 54 (1993) 1119– 1124.
Short communication
Changes in defensive behaviors following olfactory bulbectomy in male and female rats Howard S. Stock
a,b ,
*, Gregory A. Hand a , Kris Ford a , Marlene A. Wilson a
a
University of South Carolina, School of Medicine, Department of Pharmacology and Physiology, Columbia, SC 29208, USA b WJB Dorn Department of Veterans Affairs Medical Center, 6439 Garners Ferry Road, Columbia, SC 29209 -1439, USA Accepted 20 March 2001
Abstract The present study examined if olfactory bulbectomy (OBX) altered defensive behaviors on the elevated plus-maze and the open-field differently in male and female rats. Similar increases in defensive behaviors in male and female rats were observed in both tests following OBX. No significant correlations were detected between defensive behaviors and activity, supporting the hypothesis that some behavioral changes following OBX may be due to decreased defensive behaviors and not increased activity. 2001 Elsevier Science B.V. All rights reserved. Theme: Neural basis of behavior Topic: Stress Keywords: Sex difference; Defensive behavior; Plus-maze; Open-field; Model of depression
1. Introduction In stressful situations animals will elicit a variety of defensive behaviors, including: freezing, analgesia, burying, escape, and avoidance [1]. These behaviors are enhanced in novel environments and are attenuated in familiar environments. Moreover, the attenuation and enhancement of these behaviors can also be modulated by anxiolytic or anxiogenic compounds, respectively. The elevated plus-maze test, used mainly as a measure of anxiety in rodents, is based on the approach–avoidance aspect of a rat’s defensive response. It has been proposed that the open arms of the plus-maze produce a greater aversive reaction because they have a greater amount of novel stimulation compared to the closed arms (see [4] for references). Therefore, an increase in open arm time is suggestive of a decrease in a rat’s defensive responding. In addition, measuring exploration behavior the open-field test has also been used as a measure of defensive behaviors *Corresponding author. Tel.: 11-803-733-3156; fax: 11-803-7331523. E-mail address: [email protected] (H.S. Stock).
in rodents, where increased line crossings and rearing are suggestive of a decrease in defensive behaviors [14,19]. In the rat, removal of the olfactory bulbs results in a wide variety of behavioral and neurochemical changes including decreased serotonin levels, decreased norepinephrine levels, increased corticosterone levels, and increased gama-aminobutyric acid (GABA) levels [8]. Some of the behavioral changes associated with olfactory bulbectomy (OBX), such as increased open-field activity levels, decrements in passive avoidance tasks, increased time spent in open arms in the elevated plus-maze [8], decreased shock-induced freezing, and decreased time spent in a covered enclosure, are thought to be associated with a decrement in the animal’s defensive responding and not due to an increase in activity levels [11]. The majority of previous research examining OBX-induced changes in defensive behaviors have predominantly used male rats. Recently, we have observed sex and gonadal hormone effects on open-field behavior and sucrose preference tasks following OBX [16]. In addition, sex and gonadectomy effects have been observed in GABA–benzodiazepine (BZ) reactivity [20], a system important in modulating defensive responding [3,9,12,13]. Therefore, the present
0006-8993 / 01 / $ – see front matter 2001 Elsevier Science B.V. All rights reserved. PII: S0006-8993( 01 )02421-0
H.S. Stock et al. / Brain Research 903 (2001) 242 – 246
study was designed to determine if the sex and the gonadal status of the animal interact with OBX to alter defensive responding as measured by changes in elevated plus-maze or open-field related behaviors.
2. Methods
2.1. Subjects Male and female Long–Evans hooded rats (Harlan Sprague–Dawley Inc, Indianapolis, IN, USA) were housed singly with food and water available ad libitum. The animals were kept on a 12–12 h light–dark cycle, lights on at 7:00 am, with behavioral testing occurring in the early part of the light phase (0900–1200 h). Animals were housed in an AAALAC-approved animal colony room and all procedures were approved by the University of South Carolina Animal Care and Use Committee.
2.2. Handling and surgeries Handling and surgery techniques were identical to those described in Ref. [16]. Eight groups were tested: male sham (N56), male OBX (N56), orchidectomized (ORCH) sham (N56), OBX–ORCH (N57), female sham (N56), female OBX (N56), ovariectomized (OVX) sham (N55), and OBX–OVX (N55).
2.3. Apparatus and test procedure To determine the interactions between the sex, gonadal status of the animal, and OBX on behavioral responding in a novel open-field animals were tested 2 weeks following OBX in a open-field apparatus as described in Ref. [7]. The animals were placed into the center of the open-field and allowed to explore freely for 5 min. Behavior was video-taped via a video camera suspended 6 feet above the center of the maze. The video tapes were scored at a later date by an experimenter who was blind to the treatment conditions of the animals. The behaviors of interest were the number of lines crossed and the number of rears made during exposure to the open-field. A line crossing was defined as all four paws crossing over the line. Changes in the defensive behaviors following OBX were also assessed using an elevated plus-maze identical to that described in Stock et al. [16]. Twenty-four hours following open-field testing animals were placed into the center square and allowed to explore the maze for a period of 5 min [10]. Behavior was taped via a video camera suspended 3 feet above the center of the maze. The video tapes were scored at a later date by an experimenter who was blind to the treatment conditions of the animals. The behaviors of interest on the elevated plus-maze were percent open arm time (% open arm time), percent open arm entries (% open arm entries), and center square time.
243
An animal was scored as having entered the arm when all four paws were inside the arm. Percent open arm time was calculated as the total time spent in the open arms divided by the total time spent in all arms of the maze. Percent open arm entries was calculated as the total entries into the open arm divided by the total number of entries into all arms of the maze. Both percent open arm time and percent open arm entries have been reported to be behavioral indices of defensive responding in rats, with increases in time and entries suggesting lower defensive behaviors [4,10]. Center square time has been reported as a reliable measure of activity levels for this type of elevated plusmaze (i.e., ledges on open arms) [2].
2.4. Statistics The influences of sex (male vs. female), gonadectomy (intact vs. gonadectomized), and OBX condition (sham vs. OBX) on plus-maze and open-field related behaviors were compared by a three-way analysis of variance (ANOVA; sex, gonadectomy, and OBX as the between-subject factors). The three-way ANOVA with sex, gonadectomy, and OBX condition revealed no main effects or interactions of gonadectomy on any dependent measures. Consequently, data were collapsed across gonadectomy and analyzed with a two-way analysis of variance (sex and OBX condition). For all statistical analyses, Tukey honestly significant difference tests were used as post-hoc tests. The alpha level for all statistical analysis was 0.05. Pearson product moment correlations were conducted between open-field related behaviors and the behavioral measures recorded on the plus-maze.
3. Results
3.1. Number of lines crossed As seen in Fig. 1A, OBX significantly increased the number of line crossings compared to sham animals. A significant OBX main effect was detected [F(1,43)542.38, P50.001]. A Tukey post-hoc test demonstrated that OBX animals showed significantly greater number of lines crossed than sham controls (P,0.05). Pearson’s product moment correlation reported a significant positive correlation between number of lines crossed and number of rears (r 2 50.80, P50.001), percent open arm time (r 2 50.40, P50.005), and percent open arm entries (r 2 50.32 P5 0.027).
3.2. Number of rears As seen in Fig. 1B, OBX significantly increased the number of rears compared to sham animals. A significant OBX main effect was detected [F(1,43)559.17, P5 0.001]. A Tukey post-hoc test demonstrated that OBX
244
H.S. Stock et al. / Brain Research 903 (2001) 242 – 246
Fig. 1. Open-field related behaviors: (A) Each bar represents mean number of lines crossed (6S.E.M.) measured during a 5-min exposure to the open-field apparatus. Animals underwent either olfactory bulbectomy [OBX: dark columns (male N513; female N511)] or sham surgery [sham: open columns (male N512; female N511)]. *OBX animals had significantly greater number of lines crossed than sham controls (P, 0.05). (B) Each bar represents mean number of rears (6S.E.M.) measured during a 5-min exposure to the open-field apparatus. Animals underwent either olfactory bulbectomy [OBX: dark columns (male N513; female N511)] or sham surgery [sham: open columns (male N512; female N511)]. *OBX animals had significantly greater number of rears than sham controls (P,0.05).
animals showed a significantly greater number of rears than sham controls (P,0.05). Pearson’s product moment correlations reported a significant positive correlation between number of rears and percent open arm time (r 2 50.33 P50.022) and percent open arm entries (r 2 5 0.36 P50.010).
3.3. Percent open arm entries As seen in Fig. 2A, OBX significantly increased percent open arm entries compared to sham animals. A significant OBX main effect was detected [F(1,43)510.84, P5 0.002]. A Tukey post-hoc test demonstrated that OBX animals showed significantly greater percent open arm
Fig. 2. Plus-maze related behaviors: (A) Each bar represents mean percent open arm entries (6S.E.M.) measured during a 5-min exposure to the elevated plus-maze. Animals underwent either olfactory bulbectomy [OBX: dark columns (male N513; female N511)] or sham surgery [sham: open columns (male N512; female N511)]. *OBX animals had significantly greater percent open arm entries than sham controls (P, 0.05). (B) Each bar represents mean percent open arm time (6S.E.M.) measured during a 5-min exposure to the elevated plus-maze. Animals underwent either olfactory bulbectomy [OBX: dark columns (male N513; female N511)] or sham surgery [sham: open columns (male N512; female N511)]. *OBX animals had significantly greater percent open arm time than sham controls (P,0.05). (C) Each bar represents mean center square time (6S.E.M.) measured during a 5-min exposure to the elevated plus-maze. Animals underwent either olfactory bulbectomy [OBX: dark columns (male N513; female N511)] or sham surgery [sham: open columns (male N512; female N511)].
entries than sham controls (P,0.05). Pearson’s product moment correlations reported a significant positive correlation between percent open arm entries and percent open arm time (r 2 50.89 P50.001)
H.S. Stock et al. / Brain Research 903 (2001) 242 – 246
3.4. Percent open arm time As seen in Fig. 2B, OBX significantly increased percent open arm time compared to sham animals. A significant OBX main effect was detected [F(1,42)58.50, P50.005]. A Tukey post-hoc test demonstrated that OBX animals showed significantly greater percent open arm time than sham controls (P,0.05).
3.5. Center square time As seen in Fig. 2C, no effect of OBX on center square time was observed.
4. Discussion The present study was conducted to examine the possible interaction between OBX and the gonadal hormone milieu on changes in the rat’s defensive behavior, as measured by the open-field and elevated plus-maze. The results show that OBX increases the number of lines crossed as well as the number of rears in the open-field apparatus compared to sham controls, across all hormone groups. In addition, OBX increased the amount of time and entries on the open arms of the elevated plus maze compared to sham controls, across all hormone groups. The increases in number of lines crossed, rears, open arm time, and open arm entries suggest that OBX decreased defensive responding compared to sham controls. Moreover, this decrease in defensive behaviors in the elevated plus-maze does not appear to be due to changes in activity levels, since alterations in center square time, a more sensitive measure of activity with this particular type of plus-maze, was not observed [2]. Moreover, significant positive correlations were detected between open-field related behaviors (i.e., number of lines crossed and rears) and percent open arm time and entries. These findings suggest a relationship between these two tests in regards to defensive behaviors in rats. In the present study no significant correlations between center square time and number of lines crossed, rears, percent open arm time, and percent open arm entries were detected, suggesting no relationship between these measures of defensive behavior and activity. However, prior research suggests that percent open arm entries may be a more valid measure of the approach– avoidance component of the elevated plus-maze than percent open arm time, since it does not appear to be affected by changes in activity [4]. Overall, these results further support the hypothesis that OBX results in a decrease in the rat’s defensive behavior during exposure to an novel environment [11]. In addition, our findings suggest that modulation of gonadal hormone levels via adult gonadectomy does not alter defensive responding
245
following OBX as measured by the open-field and elevated plus-maze. Previous research has reported that control female rats exhibit greater levels of percent open arm time and entries than control male rats [7,21], although this is not a universal finding [5,15]. These discrepancies in outcomes may be due to a variety of procedural differences between these studies; including structural differences between the testing apparatus and strain of rat tested. Our previous research demonstrated that during exposure to an open-field only OBX-gonadectomized male and female rats exhibited a significant change from baseline 2 weeks following OBX [16]. However, in the present study all OBX animals exhibited an increase in open-field related behaviors. Differences in open-field testing procedures may provide a possible explanation for the discrepancy between these two studies. In the present study, the animals were tested in a novel open-field while in our previous study animals were habituated to the open-field before undergoing OBX surgery (i.e., baseline testing). Therefore, habituation and the gonadal status of the animal may have interacted to alter the increase in open-field related behaviors that have previously been observed following OBX [8]. This hypothesis is supported by the finding that gonadally-intact males that were habituated to an open-field apparatus failed to show an increase in open-field activity levels when tested following OBX surgery [17]. Central administration of a variety of compounds (i.e., benzodiazepines) that augment GABAergic functioning have been reported to alter a multitude of defensive behaviors such as freezing, exploration of a novel environment, food intake in a novel environment, and open arm time and entries [12,13]. In addition, prior research has demonstrated that OBX results in an increase in GABAergic functioning in a variety of brain areas (limbic system, cortex) [6,8,18]. Therefore, OBX may result in an increase in GABAergic activity, which in turn may result in a decrease in the defensive responding that has been observed in the present study and in previous research [8,11]. The results of this study replicate prior work that has demonstrated that OBX decreased defensive responding, without altering activity levels in male rats [8,11], and extends this finding to female, gonadectomized male, and ovariectomized female rats. A likely mediator of the decreased defensive responding following OBX may be increased GABAergic activity, however, further investigation will be needed to validate this hypothesis.
Acknowledgements The authors thank Cynthia Gobbel and Stefany Primeaux for helpful comments on the manuscript. This work was supported by a grant from The National Alliance for the Research on Schizophrenia and Depression.
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