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Effects of gender and gonadal hormones on nociceptive responses to intraplantar carrageenan in the rat
Neuroscience Letters 354 (2004) 239–241 www.elsevier.com/locate/neulet
Effects of gender and gonadal hormones on nociceptive responses to intraplantar carrageenan in the rat Jill M. Talla, Terriann Crispb,* b
a Department of Neurobiology and Pharmacology, Northeastern Ohio Universities College of Medicine, Rootstown, OH, USA Department of Physiology and Pharmacology, Des Moines University, 3200 Grand Avenue, Des Moines, IA 50312-4198, USA
Received 4 April 2003; received in revised form 2 September 2003; accepted 29 September 2003
Abstract The effects of gender and gonadal hormones on nociceptive responses to intraplantar carrageenan in the rat were investigated. A plantar analgesic meter was used to measure carrageenan-induced changes in paw withdrawal latency (PWL) values, and von Frey monofilaments were used to assess changes in paw withdrawal threshold (PWT) values in response to tactile-evoked (mechanical) stimuli. The data revealed that PWL values were significantly greater in gonadally-intact females than in gonadally-intact males, and this response was abolished by surgical gonadectomies. Gonadally-intact as well as gonadectomized male and female rats also exhibited significant carrageenan-induced decreases in PWT, but neither sex- nor gonadectomy-related differences were detected in the development of mechanical allodynia. These findings demonstrate that intraplantar carrageenan induces nociceptive behaviors in rats that are differentially affected by sex and gonadal hormones. q 2003 Published by Elsevier Ireland Ltd. Keywords: Estrogen; Hyperalgesia; Allodynia; Inflammation
Gender-related differences in the susceptibility of males and females to nociceptive stimuli have been reported [4,12] but the results are inconclusive [2,8]. In this study, we examined the effects of sex and gonadal hormones on carrageenaninduced thermal hyperalgesia and mechanical allodynia. We hypothesize that the hormonal status of gonadally-intact females augments the activity of the descending paininhibitory system [11]. Gender-related differences in acute inflammatory nociception were evaluated in 3– 6-month-old male and female Fischer 344 FBNF1 hybrid rats. Four different treatment cohorts were investigated: (1) gonadally-intact males; (2) gonadally-intact females; (3) castrated males; and (4) ovariectomized females. Ovariectomies or castrations were performed via a single abdominal incision along the midline following i.p. administration of pentobarbital (females 40 mg/kg; males 45 mg/kg) 10 –14 days prior to carrageenan injections. Carrageenan (0.15 ml of a 4% carrageenan suspension in sterile 0.9% NaCl) was inserted between the second and third metatarsal of the plantar * Corresponding author. Tel.: þ 1-515-271-1431; fax: þ1-515-271-7183. E-mail address: [email protected] (T. Crisp). 0304-3940/03/$ - see front matter q 2003 Published by Elsevier Ireland Ltd. doi:10.1016/j.neulet.2003.09.081
surface of the experimental hind paw during light halothane anesthesia [13]. Paw withdrawal latency (PWL) values were recorded from the left and right hind paws of rats using a plantar analgesic meter [6]. Three PWL values were recorded to the nearest 0.1 s, and a 20 s limit was imposed to preclude tissue damage. An average of the last two PWL values was calculated and defined as the mean PWL. Calibrated von Frey filaments were used to assess paw withdrawal threshold (PWT) values in response to mechanical (tactile) stimuli. The PWT was defined as the minimum gram strength eliciting two positive sequential responses (lifting of the paw). The behavioral testing regimen was as follows: pre-carrageenan baseline PWL and PWT testing ! castration or ovariectomy ! 10 –14 day interval ! measurement of paw thickness and width ! unilateral intraplantar carrageenan injection ! 2 h ! assessment of post-carrageenan paw thickness, PWL and PWT values. A second series of experiments was designed to determine if the particular stage of estrous influenced the carrageenan-induced responses of gonadally-intact females. Female rats displaying a 4 day estrous cycle [5,9] were used in this study. All data are presented as mean ^ SEM. PWL
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J.M. Tall, T. Crisp / Neuroscience Letters 354 (2004) 239–241
and PWT values represented the dependent factors in this investigation, and each was analyzed separately by an analysis of variance (statistical significance P # 0:05). Control animals were not injected with carrageenan. Data were further analyzed using post-hoc Bonferroni multiple comparison testing. The results revealed that gonadally-intact and gonadectomized male and female rats developed thermal hyperalgesia 2 h post-carrageenan injections, and significant sex-related differences were observed between gonadallyintact males and females. In both the control and carrageenan-treated groups, PWL values were greater in the gonadally-intact females than males. Further, sexrelated differences in PWL were abolished when gonadectomies were performed (Fig. 1). PWT values from carrageenan-injected hind paws were significantly less than control values in all treatment cohorts (Fig. 2). Allodynic responses were more robust in gonadally-intact males than females. Significant nociceptive responses were evoked in gonadally-intact females, regardless of the estrous phase. During proestrus, PWL values from the carrageenaninjected hind paw were significantly greater than those from females in estrus and diestrus (Fig. 3). In contrast, the estrous cycle did not significantly affect the development or magnitude of mechanical allodynia (Fig. 4). The results of these investigations demonstrated that: (1) gonadally-intact and gonadectomized male and female rats displayed thermal hyperalgesia 2 h post-carrageenan injection; (2) PWL values differed significantly between gonadally-intact male and female rats in both the control and carrageenan-treated groups; and (3) surgical gonadectomies completely abolished this effect. Spinopetal serotonergic (5-HT) and noradrenergic (NE) fibers exert
Fig. 1. PWL values from the control and carrageenan-injected hind paws of gonadally-intact and gonadectomized male and female rats. Values represent the mean PWL ^ SEM (n ¼ 9 rats/treatment group); *P # 0:05 versus sex-matched PWL controls; FP # 0:05, PWL values of gonadally-intact males versus gonadally-intact females in both the control and treatment conditions.
Fig. 2. PWT measurements from the control and carrageenan-injected hind paws of gonadally-intact and gonadectomized male and female rats. Values represent the mean PWT ^ SEM (n ¼ 9 rats/treatment group); *P # 0:05 versus sex-matched PWT controls; FP # 0:05, carrageenan-induced PWT of gonadally-intact males versus intact and ovariectomized females.
antinociceptive effects at the level of the spinal dorsal horn [1]. Monoamine oxidase (MAO) enzymatically metabolizes 5-HT and NE, and the enzymatic activity of MAO is altered by gonadal steroid hormones [10,14]. Estrogen-induced inhibition of MAO-A and MAO-B activity in the dorsal horn of the spinal cord [7] could conceivably increase local spinal biogenic amines. Thus, the descending pain-inhibitory system may play a more prominent role in modulating nociceptive input in gonadally-intact female rats. Substance P (SP) is involved in the transmission of nociceptive information at the spinal level, and ovarian hormones modulate the concentration of SP. An inverse relationship has been reported for 17 b-estradiol and SP [3];
Fig. 3. PWL values from control and carrageenan-injected hind paws of gonadally-intact female rats during proestrus, estrus and diestrus. Values represent the mean ^ SEM (n ¼ 10 rats/estrous stage); *P # 0:05 versus estrous stage-matched controls; KP # 0:05 versus carrageenan-induced estrus and diestrus PWL values.
J.M. Tall, T. Crisp / Neuroscience Letters 354 (2004) 239–241
241
References
Fig. 4. PWT measurements from the control and carrageenan-injected hind paws of gonadally-intact female rats during proestrus, estrus and diestrus. Values represent the mean ^ SEM (n ¼ 10 rats/estrous stage); *P # 0:05 versus estrous stage-matched PWT control hind paw values.
SP concentrations are lowest during the estrous stage when 17 b-estradiol levels are elevated. These investigators also reported that spinal SP levels were higher in gonadallyintact male rats compared to intact or ovariectomized females [3]. The negative effect of estrogen on SP content could blunt spinal nociceptive input. In an earlier study in our lab, gonadally-intact females were less sensitive than males to noxious thermal stimulation following peripheral nerve injury [11], supporting the hypothesis that SPmediated nociceptive transmission is attenuated by ovarian steroid hormones. In conclusion, gender and gonadal hormones influence the behavioral manifestations of carrageenan-induced nociception. The clinical implications of sex-related differences in the sensitivity to and treatment of pain remain to be elucidated. A heightened awareness of the role of sex steroids in nociceptive processing may promote the integration of hormonal factors into the intervention and management of chronic pain disorders.
[1] A.I. Basbaum, H.L. Fields, Endogenous pain control systems: brainstem spinal pathways and endorphin circuitry, Annu. Rev. Neurosci. 7 (1984) 309 –338. [2] K.J. Berkley, Sex differences in pain, Behav. Brain Sci. 20 (1997) 371–380. [3] P. Duval, V. Lenoir, S. Moussaoui, C. Garret, B. Kerdelhue´, Substance P and neurokinin A variations throughout the rat estrous cycle; comparison with ovariectomized and male rats: II. Trigeminal nucleus and cervical spinal cord, J. Neurosci. Res. 45 (1996) 610–616. [4] R.B. Fillingim, W. Maixner, Gender differences in the responses to noxious stimuli, Pain Forum 4 (1995) 209–221. [5] M.E. Freeman, The neuroendocrine control of the ovarian cycle of the rat, in: E. Knobil, J.D. Neill (Eds.), The Physiology of Reproduction, 2nd Edition., Raven Press, New York, 1994, pp. 613 –615. [6] K. Hargreaves, R. Dubner, F. Brown, C. Flores, J. Joris, A new and sensitive method for measuring thermal nociception in cutaneous hyperalgesia, Pain 32 (1988) 77–88. [7] D.P. Holschneider, T. Kumazawa, K. Chen, J.C. Shih, Tissue-specific effects of estrogen on monoamine oxidase A and B in the rat, Life Sci. 63 (1998) 155 –160. [8] K.L. Kepler, B. Kest, J.M. Kiefel, M.L. Cooper, R.J. Bodnar, Roles of gender, gonadectomy and estrous phase in the analgesic effects of intracerebroventricular morphine in rats, Pharmacol. Biochem. Behav. 34 (1989) 119–127. [9] J.A. Long, H.M. Evans, The estrous cycle of the rat and its associated phenomena, Mem. Univ. California 6 (1922) 1–148. [10] Z.-Q. Ma, E. Violani, F. Villa, G.B. Picotti, A. Maggi, Estrogenic control of monoamine oxidase A activity in human neuroblastoma cells expressing physiological concentrations of estrogen receptor, Eur. J. Pharmacol. 284 (1995) 171–176. [11] J.M. Tall, S.L. Stuesse, W.L.R. Cruce, T. Crisp, Gender and the behavioral manifestations of neuropathic pain, Pharmacol. Biochem. Behav. 68 (2001) 99–104. [12] A. Unruh, Gender variations in clinical pain experience, Pain 65 (1996) 123–167. [13] C.A. Winter, E.A. Risley, G.W. Nuss, Carrageenin-induced edema in hind paw of the rat as an assay for antiinflammatory drugs, Proc. Soc. Exp. Biol. Med. 111 (1962) 544–547. [14] A.J. Zolovick, R. Pearse, K.W. Boehlke, B.E. Eleftheriou, Monoamine oxidase activity in various parts of the rat brain during the estrous cycle, Science 154 (1966) 649.
Effects of gender and gonadal hormones on nociceptive responses to intraplantar carrageenan in the rat Jill M. Talla, Terriann Crispb,* b
a Department of Neurobiology and Pharmacology, Northeastern Ohio Universities College of Medicine, Rootstown, OH, USA Department of Physiology and Pharmacology, Des Moines University, 3200 Grand Avenue, Des Moines, IA 50312-4198, USA
Received 4 April 2003; received in revised form 2 September 2003; accepted 29 September 2003
Abstract The effects of gender and gonadal hormones on nociceptive responses to intraplantar carrageenan in the rat were investigated. A plantar analgesic meter was used to measure carrageenan-induced changes in paw withdrawal latency (PWL) values, and von Frey monofilaments were used to assess changes in paw withdrawal threshold (PWT) values in response to tactile-evoked (mechanical) stimuli. The data revealed that PWL values were significantly greater in gonadally-intact females than in gonadally-intact males, and this response was abolished by surgical gonadectomies. Gonadally-intact as well as gonadectomized male and female rats also exhibited significant carrageenan-induced decreases in PWT, but neither sex- nor gonadectomy-related differences were detected in the development of mechanical allodynia. These findings demonstrate that intraplantar carrageenan induces nociceptive behaviors in rats that are differentially affected by sex and gonadal hormones. q 2003 Published by Elsevier Ireland Ltd. Keywords: Estrogen; Hyperalgesia; Allodynia; Inflammation
Gender-related differences in the susceptibility of males and females to nociceptive stimuli have been reported [4,12] but the results are inconclusive [2,8]. In this study, we examined the effects of sex and gonadal hormones on carrageenaninduced thermal hyperalgesia and mechanical allodynia. We hypothesize that the hormonal status of gonadally-intact females augments the activity of the descending paininhibitory system [11]. Gender-related differences in acute inflammatory nociception were evaluated in 3– 6-month-old male and female Fischer 344 FBNF1 hybrid rats. Four different treatment cohorts were investigated: (1) gonadally-intact males; (2) gonadally-intact females; (3) castrated males; and (4) ovariectomized females. Ovariectomies or castrations were performed via a single abdominal incision along the midline following i.p. administration of pentobarbital (females 40 mg/kg; males 45 mg/kg) 10 –14 days prior to carrageenan injections. Carrageenan (0.15 ml of a 4% carrageenan suspension in sterile 0.9% NaCl) was inserted between the second and third metatarsal of the plantar * Corresponding author. Tel.: þ 1-515-271-1431; fax: þ1-515-271-7183. E-mail address: [email protected] (T. Crisp). 0304-3940/03/$ - see front matter q 2003 Published by Elsevier Ireland Ltd. doi:10.1016/j.neulet.2003.09.081
surface of the experimental hind paw during light halothane anesthesia [13]. Paw withdrawal latency (PWL) values were recorded from the left and right hind paws of rats using a plantar analgesic meter [6]. Three PWL values were recorded to the nearest 0.1 s, and a 20 s limit was imposed to preclude tissue damage. An average of the last two PWL values was calculated and defined as the mean PWL. Calibrated von Frey filaments were used to assess paw withdrawal threshold (PWT) values in response to mechanical (tactile) stimuli. The PWT was defined as the minimum gram strength eliciting two positive sequential responses (lifting of the paw). The behavioral testing regimen was as follows: pre-carrageenan baseline PWL and PWT testing ! castration or ovariectomy ! 10 –14 day interval ! measurement of paw thickness and width ! unilateral intraplantar carrageenan injection ! 2 h ! assessment of post-carrageenan paw thickness, PWL and PWT values. A second series of experiments was designed to determine if the particular stage of estrous influenced the carrageenan-induced responses of gonadally-intact females. Female rats displaying a 4 day estrous cycle [5,9] were used in this study. All data are presented as mean ^ SEM. PWL
240
J.M. Tall, T. Crisp / Neuroscience Letters 354 (2004) 239–241
and PWT values represented the dependent factors in this investigation, and each was analyzed separately by an analysis of variance (statistical significance P # 0:05). Control animals were not injected with carrageenan. Data were further analyzed using post-hoc Bonferroni multiple comparison testing. The results revealed that gonadally-intact and gonadectomized male and female rats developed thermal hyperalgesia 2 h post-carrageenan injections, and significant sex-related differences were observed between gonadallyintact males and females. In both the control and carrageenan-treated groups, PWL values were greater in the gonadally-intact females than males. Further, sexrelated differences in PWL were abolished when gonadectomies were performed (Fig. 1). PWT values from carrageenan-injected hind paws were significantly less than control values in all treatment cohorts (Fig. 2). Allodynic responses were more robust in gonadally-intact males than females. Significant nociceptive responses were evoked in gonadally-intact females, regardless of the estrous phase. During proestrus, PWL values from the carrageenaninjected hind paw were significantly greater than those from females in estrus and diestrus (Fig. 3). In contrast, the estrous cycle did not significantly affect the development or magnitude of mechanical allodynia (Fig. 4). The results of these investigations demonstrated that: (1) gonadally-intact and gonadectomized male and female rats displayed thermal hyperalgesia 2 h post-carrageenan injection; (2) PWL values differed significantly between gonadally-intact male and female rats in both the control and carrageenan-treated groups; and (3) surgical gonadectomies completely abolished this effect. Spinopetal serotonergic (5-HT) and noradrenergic (NE) fibers exert
Fig. 1. PWL values from the control and carrageenan-injected hind paws of gonadally-intact and gonadectomized male and female rats. Values represent the mean PWL ^ SEM (n ¼ 9 rats/treatment group); *P # 0:05 versus sex-matched PWL controls; FP # 0:05, PWL values of gonadally-intact males versus gonadally-intact females in both the control and treatment conditions.
Fig. 2. PWT measurements from the control and carrageenan-injected hind paws of gonadally-intact and gonadectomized male and female rats. Values represent the mean PWT ^ SEM (n ¼ 9 rats/treatment group); *P # 0:05 versus sex-matched PWT controls; FP # 0:05, carrageenan-induced PWT of gonadally-intact males versus intact and ovariectomized females.
antinociceptive effects at the level of the spinal dorsal horn [1]. Monoamine oxidase (MAO) enzymatically metabolizes 5-HT and NE, and the enzymatic activity of MAO is altered by gonadal steroid hormones [10,14]. Estrogen-induced inhibition of MAO-A and MAO-B activity in the dorsal horn of the spinal cord [7] could conceivably increase local spinal biogenic amines. Thus, the descending pain-inhibitory system may play a more prominent role in modulating nociceptive input in gonadally-intact female rats. Substance P (SP) is involved in the transmission of nociceptive information at the spinal level, and ovarian hormones modulate the concentration of SP. An inverse relationship has been reported for 17 b-estradiol and SP [3];
Fig. 3. PWL values from control and carrageenan-injected hind paws of gonadally-intact female rats during proestrus, estrus and diestrus. Values represent the mean ^ SEM (n ¼ 10 rats/estrous stage); *P # 0:05 versus estrous stage-matched controls; KP # 0:05 versus carrageenan-induced estrus and diestrus PWL values.
J.M. Tall, T. Crisp / Neuroscience Letters 354 (2004) 239–241
241
References
Fig. 4. PWT measurements from the control and carrageenan-injected hind paws of gonadally-intact female rats during proestrus, estrus and diestrus. Values represent the mean ^ SEM (n ¼ 10 rats/estrous stage); *P # 0:05 versus estrous stage-matched PWT control hind paw values.
SP concentrations are lowest during the estrous stage when 17 b-estradiol levels are elevated. These investigators also reported that spinal SP levels were higher in gonadallyintact male rats compared to intact or ovariectomized females [3]. The negative effect of estrogen on SP content could blunt spinal nociceptive input. In an earlier study in our lab, gonadally-intact females were less sensitive than males to noxious thermal stimulation following peripheral nerve injury [11], supporting the hypothesis that SPmediated nociceptive transmission is attenuated by ovarian steroid hormones. In conclusion, gender and gonadal hormones influence the behavioral manifestations of carrageenan-induced nociception. The clinical implications of sex-related differences in the sensitivity to and treatment of pain remain to be elucidated. A heightened awareness of the role of sex steroids in nociceptive processing may promote the integration of hormonal factors into the intervention and management of chronic pain disorders.
[1] A.I. Basbaum, H.L. Fields, Endogenous pain control systems: brainstem spinal pathways and endorphin circuitry, Annu. Rev. Neurosci. 7 (1984) 309 –338. [2] K.J. Berkley, Sex differences in pain, Behav. Brain Sci. 20 (1997) 371–380. [3] P. Duval, V. Lenoir, S. Moussaoui, C. Garret, B. Kerdelhue´, Substance P and neurokinin A variations throughout the rat estrous cycle; comparison with ovariectomized and male rats: II. Trigeminal nucleus and cervical spinal cord, J. Neurosci. Res. 45 (1996) 610–616. [4] R.B. Fillingim, W. Maixner, Gender differences in the responses to noxious stimuli, Pain Forum 4 (1995) 209–221. [5] M.E. Freeman, The neuroendocrine control of the ovarian cycle of the rat, in: E. Knobil, J.D. Neill (Eds.), The Physiology of Reproduction, 2nd Edition., Raven Press, New York, 1994, pp. 613 –615. [6] K. Hargreaves, R. Dubner, F. Brown, C. Flores, J. Joris, A new and sensitive method for measuring thermal nociception in cutaneous hyperalgesia, Pain 32 (1988) 77–88. [7] D.P. Holschneider, T. Kumazawa, K. Chen, J.C. Shih, Tissue-specific effects of estrogen on monoamine oxidase A and B in the rat, Life Sci. 63 (1998) 155 –160. [8] K.L. Kepler, B. Kest, J.M. Kiefel, M.L. Cooper, R.J. Bodnar, Roles of gender, gonadectomy and estrous phase in the analgesic effects of intracerebroventricular morphine in rats, Pharmacol. Biochem. Behav. 34 (1989) 119–127. [9] J.A. Long, H.M. Evans, The estrous cycle of the rat and its associated phenomena, Mem. Univ. California 6 (1922) 1–148. [10] Z.-Q. Ma, E. Violani, F. Villa, G.B. Picotti, A. Maggi, Estrogenic control of monoamine oxidase A activity in human neuroblastoma cells expressing physiological concentrations of estrogen receptor, Eur. J. Pharmacol. 284 (1995) 171–176. [11] J.M. Tall, S.L. Stuesse, W.L.R. Cruce, T. Crisp, Gender and the behavioral manifestations of neuropathic pain, Pharmacol. Biochem. Behav. 68 (2001) 99–104. [12] A. Unruh, Gender variations in clinical pain experience, Pain 65 (1996) 123–167. [13] C.A. Winter, E.A. Risley, G.W. Nuss, Carrageenin-induced edema in hind paw of the rat as an assay for antiinflammatory drugs, Proc. Soc. Exp. Biol. Med. 111 (1962) 544–547. [14] A.J. Zolovick, R. Pearse, K.W. Boehlke, B.E. Eleftheriou, Monoamine oxidase activity in various parts of the rat brain during the estrous cycle, Science 154 (1966) 649.