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Effects of age on behavioral signs of neuropathic pain in an experimental rat model
ELSEVIER
Neuroscience Letters 183 (1995) 54-57
NtllllO~lE LETTERS
Effects of age on behavioral signs of neuropathic pain in an experimental rat model J i n M o C h u n g a,b,c,*, Y o o n C h o i a, Y o u n g W o o k Y o o n a, H e u n g S i k N a a aMarine Biomedical lnstitute, Universityof Texas Medical Branch, Galveston, TX 77555-0843, USA bDepartment of Anatomy and Neurosciences, Universityof Texas Medical Branch, Galveston, TX 77555-0843, USA CDepartment of Physiology and Biophysics, Universityof Texas Medical Branch, Galveston, TX 77555-0843, USA Received 15 August 1994; revised version received 26 October 1994; accepted 26 October 1994
Abstraet The present study examined the effect that aging has on the manifestations of pain behaviors in a rat model of neuropathic pain. Two experiments were conducted. The first experiment compared young, mature and old rats. After tight ligation of the L5 and L6 segmental spinal nerves, young rats displayed much more vigorous behavioral signs of mechanical allodynia and ongoing pain than did either mature or old rats. The second experiment was done using the same rats at two different time points. Spinal nerve ligation was done on the left side of rats when they were young and the same surgery was repeated on the right side 20 weeks later (mature age). Comparison of pain behaviors between the left and right sides confirmed the results of the first experiment. The data suggest that younger rats show much more robust behavioral signs of neuropathic pain compared to older rats.
Keywords: Causalgia; Hyperalgesia; Peripheral nerve injury; Sympathetically maintained pain
We recently developed a rat model of peripheral neuropathic pain [2,7]. The model was produced by tightly ligating the L5 and L6 segmentai spinal nerves unilaterally at a site just distal to the dorsal root ganglion (DRG). With this model, we began to examine various pathophysiological mechanisms that are potentially involved in neuropathic pain [6,8,10,11]. Ail of the above studies were done using young adult rats weighing about 150--200 g. Young animais were chosen because there has been an anecdotai belief in our laboratory that older rats do not show as robust behavioral signs of neuropathic pain as do younger rats after the same peripherai nerve injury. In addition, there was a report indicating that the age and size of rats affect the development of ailodynia in another neuropathic pain model [12]. The present study systematically examined the difference in behavioral manifestations between different age groups in our rat neuropathic pain model. A total of 51 maie Sprague-Dawley rats was used. A unilateral peripheral neuropathy was induced according to * Corresponding author, Marine Biomedical lnstitute, University of Texas Medieal Braneh, 301 University Boulevard, Galveston, TX 77555-0843, USA. Tel.: +1 409 772 2106; Fax: +1 409 762 9382.
0304-3940/95/$09.50
the method previously described in detail [2,7]. Briefly, under gaseous anesthesia with a mixture of halothane (2% for induction and 0.8% for maintenance) and a 1:2 flow ratio of N20 and 02, the left L5 and L6 spinal nerves were isolated and tightly ligated with 6--0 silk thread. Hemostasis was confirmed and the wound was then sutured closed. At the end of the experiment, rats were sacrificed by an injection of an overdose of sodium pentobarbital (100 mg/kg, i.p.) followed by opening of the chest. Behavioral signs representing two different components (mechanicai ailodynia and ongoing pain) of neuropathic pain were examined as previously described in detail [2,7,10,11]. To quantify mechanical sensitivity of the foot, brisk foot withdrawal in response to normally innocuous mechanical stimuli was measured. Innocuous mechanical stimuli were applied with von Frey filaments of two different bending forces (8.4 mN (0.9 g) and 54.4 mN (5.5 g)). The rat was placed under a transparent plastic dome on a metai mesh floor and the von Frey filaments were applied to che plantar surface of the foot. A von Frey filament was applied 10 times (once every 34 s) to each hind paw. The frequency of foot withdrawal was expressed as a percentage of trials. A significant in-
© 1995 Elsevier Science Ireland Ltd. Ail rights reserved
SSDI 0304-3940(94)11113-T
J.M, Chung et al. I Neuroscience Letters 183 (1995) 54-57 crease in the frequency of brisk foot withdrawals in response to these innocuous mechanical stimuli was interpreted as mechanical «dlodynia. Two different tesa; for ongoing pain were employed: cold stress-induced ongoing pain and spontaneous pain. For the measurement o f the cold-stress induced ongoing pain, each rat was placed on a brass plate kept at a cold temperature (5 _+ 1°C) and covered by a transparent plastic dome (11 x 11 x 26 cm). After 5 min of adaptation, the eumulative duration of rime that the rat held its foot off tho floor for the next 5 min was recorded. In a previous study, we presenttfl evidence suggesting that this test reflects ongoing pain induced by cold stress [2]. For the measurement of spontaneous pain (ongoing pain without apparent external stimulus), exactly the saine procedure was used, except that the brass plate was set at a neutral temperature (30_+ I°C). Then the same footlifting measurements were done at the saine rime points as described above for cold stress-induced ongoing pain. Data are expressed as mean _+ standard error of mean (SEM). Statistical treatments were made using the Kruskal-Wallis one-way analysis of variance or the Wilcoxon matched-pairs signed-ranks test (non-parametric tests). P values less than 0.05 were considered to be significant. Two series of experiments were conducted. The first experiment of the study compared young, mature and old rats for their behavioral manifestations after the same peripheral nerve injury. To do this, batches of 40-day-old maie rats were obtained from Harlan Sprague Dawley, Inc. Neuropathic surgery was done on half of these rats (young age group; n =-- 16) when they were between 40 and 5 0 d a y s old (bod!y weight 141.4_+ 3 . 2 g (mean _+ SEM) and the other half (mature age group; n = 16) were kept until they were between 100 and 120 days old (body weight 390.8 -+ 7.9 g), at which point neuropathic surgery was donc. As the third group (old age group; n = 11), the same operation was done on a batch of 15-month-old maie rats (body weight 554.3 -+ 8.1 g) obtained from the same source. Behavioral tests were donc on these three groups of rats until 2 weeks postoperatively. Fig. 1 shows the results of this experiment. The results of the behavioral tests for mechanical sensitivity of the paw are shown in Fig. lA,B. Before surgery, the ratg from any group were rarely responsive even to the stronger von Frey filament (Fig. lB; 54.4 mN) applied to the plantar surface of the foot. After ligation of the L5 and L6 spinal nerves, the ipsilateral hind paw became sensitive to mechanical stimuli, even with the weaker von Frey filament (Fig. lA; 8.4 mN). As has been donc previously [2,6-8,10,11], such an increase in mechanical sensitivity was interpreted as a behavioral sign of mechanical allodynia induced by peripheral nerve injury. The increase in mechanical sensitivity was much greater in the young rat group compared to the other two. The contralateral hind paw remained insensitive throughout the test period. The
55
data suggest that neuropathic injury produces much more profound behavioral signs of mechanical allodynia in young rats. The results of the behavioral tests of foot lifting on temperature-controlled plates are shown in Fig. 1C,D. Fig. 1C demonstrates that young rats lifted the foot off the neutral temperature plate (30°C) on the spinal nerveligated side for longer periods than did the older rats. In fact, the responses of mature and old rats to stimulation of the foot on the lesioned side were indistinguishable from those of the contralateral foot. Similarly, when the rats were placed on a cold plate (5°C), young rats withdrew the foot on the ligated side for significantly longer durations compared to either mature or old rats (Fig. 1D). These data suggest that ligation of the spinal nerves of 54.4 mN
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Fig. 1. Comparison of three groups of young, mature and old rats (n = 16, 16 and 11, respectively). In (A) and (B), the frequencies Of foot withdrawals to repeated me~hanical stimuli applied to the hind paw with von Frey filaments are expressed as percent (mean _. SEM) of trials; (A) and (B) are results obtained W'ithtwo different strengths of von Frey filaments of 8.4 and 54.4 mN, respectively. In (C) and (D), rats were placed on a brass plate ata n.eutral (30°C) (C) or a cold (5°C) temperature (D) and the cumulative durations of foot lifts off the fioor were measured dufing a 5 minute observation period. Postoperative rime is expressed as P for the p/¢0perative test period or as days after the spinal nerve iigations.'R6sults of young (Y), mature (M) and old (O) rat groups are shown as cirdes, square and triangles, respeefively. Results of control (Con.)afldexperiment (Exp) sides are shown as open and filled symbolS. Astefisks indicate values significantly different from corresp0nding values of the young rat group (P < 0.05 by the KruskaI-Wallis one-way analysis of variance).
J.M. Chung et al. I Neuroscience Letters 183 (1995) 54-57
56
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POSTOPERATIVE TIME (DAYS) Fig. 2. Comparison of the saine group of rats at young and mature ages (n = 8). The arrangement of the graph is the saine as in Fig. 1. The L5 and I.,6 spinal nerves were ligated on the left side 1 day after preoperatire tests when the rats were young (Y) and the responses of the left
foot are shown as eircles.The operationwas repeatedon the right side 20 weeks later (M for mature) and the responsesof the right foot after the second surgery are shown as tdangles. Asterisks indicate values signifieantly different from corresponding values at the young age (P < 0.05 by the Wilcoxonmatched-palrssignedranks test). young rats causes the affected hindlimb to be held off the temperature-controlled plates for long periods of rime, whereas older rats show much weaker behavior. In a previous study [2], we provided a rationale for our interpretation of the increased duration of foot lifting off temperature-controlled plates as a sign of ongoing pain. The data suggest that neuropathic injury produces much more profound behavioral signs of ongoing pain in young rats. The results of the above experiment suggest that a peripheral nerve injury does not induce neuropathic pain behaviors in older rat groups as robustly as in a young rat group. However, comparison of different groups makes it difficult to rule out completely the possibility that the difference is due to individual variations. Therefore, the experiment was repeated using the same rats at two different time points. Spinal nerve ligation was donc on the left side of a group of 8 rats when they were young (between 45 and 50days old (body weight 148.9_+ 5.4 g)). Twenty weeks later (at mature age), the saine surgical procedure was repeated on the right side (body weight 430.9 _+ 16.9 g). Then, comparisons were made
between the behaviors obtained on the left and right sides after the first and second operations, respectively. Fig. 2 shows the results of behavioral tests after the two operations. The results are basically the same as in the group comparison (Fig. 1). That is, spinal nerve ligation donc at the mature aged period produced a much smaller magnitude of neuropathic pain behaviors than did the saine procedure performed at the younger age. The data of the present study confirm our anecdotal belief that older rats do not show as robust behaviorai signs of neuropathic pain as do younger rats. The results suggest that behaviors following spinal nerve ligation are very different from autotomy which frequently occurs after deafferentation, since (1) ligations of spinal nerves at two segments (L5 and L6) do not produce autotomy behavior [7], and (2) incidents of autotomy following sciatic nerve transection are found to be much lower in young rats [13]. However, using the sciatic nerve chronic constriction model developed by Bennett and Xie [1], Tanck et al. [12] reported that behavioral signs of heat hyperalgesia as well as cold and mechanical allodynia were much less pronounced in older rats. They speculated that the effect may be due to differences in the size and structural integrity of the nerves between young and old rats. The present findings extend the previous report in that the influence of age in neuropathic pain behaviors includes signs of ongoing pain, In fact, the most dramatic difference between old and young animais was found in the signs of ongoing pain. There is no clear explanation for the effect of age on neuropathic pain behaviors at the present rime. However, it is likely due to a mechanism that involves other than anatomical differences in the peripheral nerve, as suggested previously [12], since the present study used tight ligation of the peripheral nerve fibers. There seem to be differences between young and old rats that affect fundamentally important steps in neuropathic pain generation mechanisms. Neuropathic pain is believed to be dependent on both central and peripheral mechanisms: sensitization of spinal dorsal horn cells maintained by ectopic discharges generated in the periphery. Therefore, age may influence one or more of the steps involved in these mechanisms. Potential mechanisms that may be influenced by age include: (1) the generator mechanism of ectopic discharges in the injured peripheral nerve (at the neuroma or DRG) [5]; (2) the degree of sprouting of sympathetic postganglionic fibers into the injury site and DRG [3,9]; or (3) the effectiveness of spinal dorsal horn sensitization. The last possibility is supported by a recent report of a decline in spinal opioid-induced antinociception as a function of age [4]. Since neuropathic pain is common in older people but older rats display much weaker behaviors, one might question the validity of rat spinal nerve ligation as an exact model for human neuropathic pain. Our interpretation, however, is that the behavioral observations in older rats
J.M. Chung et al. I Neuroscience Letters 183 (1995) 54-57
typify rat neuropathic pain whereas those in i m m a t u r e ( y o u n g ) rats are exaggerated. This interpretation arises from the fact that the m a g n i t u d e s of the observed behaviors saturate as the a n i m a i s get older so that the responses in mature (3--4 m o n t h s old) a n i m a i s are similar to those in aged (15 m o n t h s old) rats. Similar changes b e t w e e n immature and m a t u r e rats have been observed in another neuropathic pain model [12]. Nevertheless, as a practical point, the use o f y o u n g rats seems to be advantageous in most experimental conditions unless an u n u s u a l mechanism is i n v o l v e d in the exaggerated behaviors o f i m m a ture animais. This work was supported by N I H Grants NS 31680 and NS 11255. W e wish to thank Margie W a t s o n for editorial help. [1] Bennett, G.J. and Xie, Y.-K., A peripheral mononeuropathy in rat that produees disorders of pain sensation like those seen in man, Pain, 33 (1988) 87-107. [2] Choi, Y., Yoon, Y.W., Na, H.S., Kim, S.H. and Chung, J.M., Behaviorai signs of ongoing pain and cold ailodynia in a rat model of neuropathie pain, Pain, (1994) in press. [3] Chung, K., Kim, H.J., Na, H.S., Park, M.J. and Chung, J.M., Abnormalities of symp~~thetie innervation of the area of injured peripheral nerve in a rat model of neuropathie pain, Neurosei. Lett., 162 (1993) 85-88. [4] Cdsp, T., Stafinsky, J.L., Hoskins, D.L., Perni, V.C., Uram, M. and Gordon, T.L., Age-Jelated changes in the spinal antinocieep-
[5I
[6]
[7]
[8]
[9]
[10]
[1 I]
[12]
[13]
57
tive effects of DAGO, DPDPE and fl-endorphin in the rat, Brain Res., 643 (1994) 282-286. Devor, M., Jänig, W. and Michaelis, M., Modulation of activity in dorsal root ganglion neurons by sympathetie activation in nerveinjured rats, J. Neurophysiol., 71 (1994) 38-47. Kim, S.H. and Chung, J.M., Sympathectomy aIleviates mechanical allodynia in an experimental animal model for neuropathy in the rat, Neurosci. Lett., 134 (1991) 131-134. Kim, S.H. and Chang, J.M., An expefimental model for pefipheral neuropathy produced by segmental spinal nerve ligation in the rat, Pain, 50 (1992) 355-363. Kim, S.H., Na, H.S., Sh¢en, K. and Chung, J.M., Effects of sympathectomy on a rat model of peripheral neuropathy, Pain, 55 (1993) 85-92. McLachlan, E.M., Jänig, W., Devor, M. and Michaelis, M., Pefipheral nerve injury triggers noradrenergic sprouting within dorsal foot ganglia, Nature, 363 (1993) 543-546. Na, H.S., Leem, J.W. and Chung, J.M., Abnormalifies of mechanoreceptors in a rat model of neuropathie pain: possible involvement in mediating mechanical allodynia, J. Neurophysiol., 70 (1993) 522-528. Sheen, K. and Chung,,J.M., Signs of neuropathic pain depend on signais from injured nerve fibers in a rat modei, Brain Res., 610 (1993) 62-68. Tanck, E.N., Kroin, J.S., McCarthy, R.J., Penn, R.D. and Ivankovich, A.D., Effects of age and size on development of allodynia in a chronic pain model produc¢d by sciatic nerve ligation in rats, Pain, 51 (1992) 313-316. Wall, P.D., Devor, M., Inbal, R., Scadding, J.W., Schonfeld, D., Seltzer, Z. and Tomkiewicz, M.M., Autotomy following pefipherai nerve lesions: expefimental anaesthesia dolorosa, Pain, 7 (1979) 103-113.
Neuroscience Letters 183 (1995) 54-57
NtllllO~lE LETTERS
Effects of age on behavioral signs of neuropathic pain in an experimental rat model J i n M o C h u n g a,b,c,*, Y o o n C h o i a, Y o u n g W o o k Y o o n a, H e u n g S i k N a a aMarine Biomedical lnstitute, Universityof Texas Medical Branch, Galveston, TX 77555-0843, USA bDepartment of Anatomy and Neurosciences, Universityof Texas Medical Branch, Galveston, TX 77555-0843, USA CDepartment of Physiology and Biophysics, Universityof Texas Medical Branch, Galveston, TX 77555-0843, USA Received 15 August 1994; revised version received 26 October 1994; accepted 26 October 1994
Abstraet The present study examined the effect that aging has on the manifestations of pain behaviors in a rat model of neuropathic pain. Two experiments were conducted. The first experiment compared young, mature and old rats. After tight ligation of the L5 and L6 segmental spinal nerves, young rats displayed much more vigorous behavioral signs of mechanical allodynia and ongoing pain than did either mature or old rats. The second experiment was done using the same rats at two different time points. Spinal nerve ligation was done on the left side of rats when they were young and the same surgery was repeated on the right side 20 weeks later (mature age). Comparison of pain behaviors between the left and right sides confirmed the results of the first experiment. The data suggest that younger rats show much more robust behavioral signs of neuropathic pain compared to older rats.
Keywords: Causalgia; Hyperalgesia; Peripheral nerve injury; Sympathetically maintained pain
We recently developed a rat model of peripheral neuropathic pain [2,7]. The model was produced by tightly ligating the L5 and L6 segmentai spinal nerves unilaterally at a site just distal to the dorsal root ganglion (DRG). With this model, we began to examine various pathophysiological mechanisms that are potentially involved in neuropathic pain [6,8,10,11]. Ail of the above studies were done using young adult rats weighing about 150--200 g. Young animais were chosen because there has been an anecdotai belief in our laboratory that older rats do not show as robust behavioral signs of neuropathic pain as do younger rats after the same peripherai nerve injury. In addition, there was a report indicating that the age and size of rats affect the development of ailodynia in another neuropathic pain model [12]. The present study systematically examined the difference in behavioral manifestations between different age groups in our rat neuropathic pain model. A total of 51 maie Sprague-Dawley rats was used. A unilateral peripheral neuropathy was induced according to * Corresponding author, Marine Biomedical lnstitute, University of Texas Medieal Braneh, 301 University Boulevard, Galveston, TX 77555-0843, USA. Tel.: +1 409 772 2106; Fax: +1 409 762 9382.
0304-3940/95/$09.50
the method previously described in detail [2,7]. Briefly, under gaseous anesthesia with a mixture of halothane (2% for induction and 0.8% for maintenance) and a 1:2 flow ratio of N20 and 02, the left L5 and L6 spinal nerves were isolated and tightly ligated with 6--0 silk thread. Hemostasis was confirmed and the wound was then sutured closed. At the end of the experiment, rats were sacrificed by an injection of an overdose of sodium pentobarbital (100 mg/kg, i.p.) followed by opening of the chest. Behavioral signs representing two different components (mechanicai ailodynia and ongoing pain) of neuropathic pain were examined as previously described in detail [2,7,10,11]. To quantify mechanical sensitivity of the foot, brisk foot withdrawal in response to normally innocuous mechanical stimuli was measured. Innocuous mechanical stimuli were applied with von Frey filaments of two different bending forces (8.4 mN (0.9 g) and 54.4 mN (5.5 g)). The rat was placed under a transparent plastic dome on a metai mesh floor and the von Frey filaments were applied to che plantar surface of the foot. A von Frey filament was applied 10 times (once every 34 s) to each hind paw. The frequency of foot withdrawal was expressed as a percentage of trials. A significant in-
© 1995 Elsevier Science Ireland Ltd. Ail rights reserved
SSDI 0304-3940(94)11113-T
J.M, Chung et al. I Neuroscience Letters 183 (1995) 54-57 crease in the frequency of brisk foot withdrawals in response to these innocuous mechanical stimuli was interpreted as mechanical «dlodynia. Two different tesa; for ongoing pain were employed: cold stress-induced ongoing pain and spontaneous pain. For the measurement o f the cold-stress induced ongoing pain, each rat was placed on a brass plate kept at a cold temperature (5 _+ 1°C) and covered by a transparent plastic dome (11 x 11 x 26 cm). After 5 min of adaptation, the eumulative duration of rime that the rat held its foot off tho floor for the next 5 min was recorded. In a previous study, we presenttfl evidence suggesting that this test reflects ongoing pain induced by cold stress [2]. For the measurement of spontaneous pain (ongoing pain without apparent external stimulus), exactly the saine procedure was used, except that the brass plate was set at a neutral temperature (30_+ I°C). Then the same footlifting measurements were done at the saine rime points as described above for cold stress-induced ongoing pain. Data are expressed as mean _+ standard error of mean (SEM). Statistical treatments were made using the Kruskal-Wallis one-way analysis of variance or the Wilcoxon matched-pairs signed-ranks test (non-parametric tests). P values less than 0.05 were considered to be significant. Two series of experiments were conducted. The first experiment of the study compared young, mature and old rats for their behavioral manifestations after the same peripheral nerve injury. To do this, batches of 40-day-old maie rats were obtained from Harlan Sprague Dawley, Inc. Neuropathic surgery was done on half of these rats (young age group; n =-- 16) when they were between 40 and 5 0 d a y s old (bod!y weight 141.4_+ 3 . 2 g (mean _+ SEM) and the other half (mature age group; n = 16) were kept until they were between 100 and 120 days old (body weight 390.8 -+ 7.9 g), at which point neuropathic surgery was donc. As the third group (old age group; n = 11), the same operation was done on a batch of 15-month-old maie rats (body weight 554.3 -+ 8.1 g) obtained from the same source. Behavioral tests were donc on these three groups of rats until 2 weeks postoperatively. Fig. 1 shows the results of this experiment. The results of the behavioral tests for mechanical sensitivity of the paw are shown in Fig. lA,B. Before surgery, the ratg from any group were rarely responsive even to the stronger von Frey filament (Fig. lB; 54.4 mN) applied to the plantar surface of the foot. After ligation of the L5 and L6 spinal nerves, the ipsilateral hind paw became sensitive to mechanical stimuli, even with the weaker von Frey filament (Fig. lA; 8.4 mN). As has been donc previously [2,6-8,10,11], such an increase in mechanical sensitivity was interpreted as a behavioral sign of mechanical allodynia induced by peripheral nerve injury. The increase in mechanical sensitivity was much greater in the young rat group compared to the other two. The contralateral hind paw remained insensitive throughout the test period. The
55
data suggest that neuropathic injury produces much more profound behavioral signs of mechanical allodynia in young rats. The results of the behavioral tests of foot lifting on temperature-controlled plates are shown in Fig. 1C,D. Fig. 1C demonstrates that young rats lifted the foot off the neutral temperature plate (30°C) on the spinal nerveligated side for longer periods than did the older rats. In fact, the responses of mature and old rats to stimulation of the foot on the lesioned side were indistinguishable from those of the contralateral foot. Similarly, when the rats were placed on a cold plate (5°C), young rats withdrew the foot on the ligated side for significantly longer durations compared to either mature or old rats (Fig. 1D). These data suggest that ligation of the spinal nerves of 54.4 mN
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Fig. 1. Comparison of three groups of young, mature and old rats (n = 16, 16 and 11, respectively). In (A) and (B), the frequencies Of foot withdrawals to repeated me~hanical stimuli applied to the hind paw with von Frey filaments are expressed as percent (mean _. SEM) of trials; (A) and (B) are results obtained W'ithtwo different strengths of von Frey filaments of 8.4 and 54.4 mN, respectively. In (C) and (D), rats were placed on a brass plate ata n.eutral (30°C) (C) or a cold (5°C) temperature (D) and the cumulative durations of foot lifts off the fioor were measured dufing a 5 minute observation period. Postoperative rime is expressed as P for the p/¢0perative test period or as days after the spinal nerve iigations.'R6sults of young (Y), mature (M) and old (O) rat groups are shown as cirdes, square and triangles, respeefively. Results of control (Con.)afldexperiment (Exp) sides are shown as open and filled symbolS. Astefisks indicate values significantly different from corresp0nding values of the young rat group (P < 0.05 by the KruskaI-Wallis one-way analysis of variance).
J.M. Chung et al. I Neuroscience Letters 183 (1995) 54-57
56
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POSTOPERATIVE TIME (DAYS) Fig. 2. Comparison of the saine group of rats at young and mature ages (n = 8). The arrangement of the graph is the saine as in Fig. 1. The L5 and I.,6 spinal nerves were ligated on the left side 1 day after preoperatire tests when the rats were young (Y) and the responses of the left
foot are shown as eircles.The operationwas repeatedon the right side 20 weeks later (M for mature) and the responsesof the right foot after the second surgery are shown as tdangles. Asterisks indicate values signifieantly different from corresponding values at the young age (P < 0.05 by the Wilcoxonmatched-palrssignedranks test). young rats causes the affected hindlimb to be held off the temperature-controlled plates for long periods of rime, whereas older rats show much weaker behavior. In a previous study [2], we provided a rationale for our interpretation of the increased duration of foot lifting off temperature-controlled plates as a sign of ongoing pain. The data suggest that neuropathic injury produces much more profound behavioral signs of ongoing pain in young rats. The results of the above experiment suggest that a peripheral nerve injury does not induce neuropathic pain behaviors in older rat groups as robustly as in a young rat group. However, comparison of different groups makes it difficult to rule out completely the possibility that the difference is due to individual variations. Therefore, the experiment was repeated using the same rats at two different time points. Spinal nerve ligation was donc on the left side of a group of 8 rats when they were young (between 45 and 50days old (body weight 148.9_+ 5.4 g)). Twenty weeks later (at mature age), the saine surgical procedure was repeated on the right side (body weight 430.9 _+ 16.9 g). Then, comparisons were made
between the behaviors obtained on the left and right sides after the first and second operations, respectively. Fig. 2 shows the results of behavioral tests after the two operations. The results are basically the same as in the group comparison (Fig. 1). That is, spinal nerve ligation donc at the mature aged period produced a much smaller magnitude of neuropathic pain behaviors than did the saine procedure performed at the younger age. The data of the present study confirm our anecdotal belief that older rats do not show as robust behaviorai signs of neuropathic pain as do younger rats. The results suggest that behaviors following spinal nerve ligation are very different from autotomy which frequently occurs after deafferentation, since (1) ligations of spinal nerves at two segments (L5 and L6) do not produce autotomy behavior [7], and (2) incidents of autotomy following sciatic nerve transection are found to be much lower in young rats [13]. However, using the sciatic nerve chronic constriction model developed by Bennett and Xie [1], Tanck et al. [12] reported that behavioral signs of heat hyperalgesia as well as cold and mechanical allodynia were much less pronounced in older rats. They speculated that the effect may be due to differences in the size and structural integrity of the nerves between young and old rats. The present findings extend the previous report in that the influence of age in neuropathic pain behaviors includes signs of ongoing pain, In fact, the most dramatic difference between old and young animais was found in the signs of ongoing pain. There is no clear explanation for the effect of age on neuropathic pain behaviors at the present rime. However, it is likely due to a mechanism that involves other than anatomical differences in the peripheral nerve, as suggested previously [12], since the present study used tight ligation of the peripheral nerve fibers. There seem to be differences between young and old rats that affect fundamentally important steps in neuropathic pain generation mechanisms. Neuropathic pain is believed to be dependent on both central and peripheral mechanisms: sensitization of spinal dorsal horn cells maintained by ectopic discharges generated in the periphery. Therefore, age may influence one or more of the steps involved in these mechanisms. Potential mechanisms that may be influenced by age include: (1) the generator mechanism of ectopic discharges in the injured peripheral nerve (at the neuroma or DRG) [5]; (2) the degree of sprouting of sympathetic postganglionic fibers into the injury site and DRG [3,9]; or (3) the effectiveness of spinal dorsal horn sensitization. The last possibility is supported by a recent report of a decline in spinal opioid-induced antinociception as a function of age [4]. Since neuropathic pain is common in older people but older rats display much weaker behaviors, one might question the validity of rat spinal nerve ligation as an exact model for human neuropathic pain. Our interpretation, however, is that the behavioral observations in older rats
J.M. Chung et al. I Neuroscience Letters 183 (1995) 54-57
typify rat neuropathic pain whereas those in i m m a t u r e ( y o u n g ) rats are exaggerated. This interpretation arises from the fact that the m a g n i t u d e s of the observed behaviors saturate as the a n i m a i s get older so that the responses in mature (3--4 m o n t h s old) a n i m a i s are similar to those in aged (15 m o n t h s old) rats. Similar changes b e t w e e n immature and m a t u r e rats have been observed in another neuropathic pain model [12]. Nevertheless, as a practical point, the use o f y o u n g rats seems to be advantageous in most experimental conditions unless an u n u s u a l mechanism is i n v o l v e d in the exaggerated behaviors o f i m m a ture animais. This work was supported by N I H Grants NS 31680 and NS 11255. W e wish to thank Margie W a t s o n for editorial help. [1] Bennett, G.J. and Xie, Y.-K., A peripheral mononeuropathy in rat that produees disorders of pain sensation like those seen in man, Pain, 33 (1988) 87-107. [2] Choi, Y., Yoon, Y.W., Na, H.S., Kim, S.H. and Chung, J.M., Behaviorai signs of ongoing pain and cold ailodynia in a rat model of neuropathie pain, Pain, (1994) in press. [3] Chung, K., Kim, H.J., Na, H.S., Park, M.J. and Chung, J.M., Abnormalities of symp~~thetie innervation of the area of injured peripheral nerve in a rat model of neuropathie pain, Neurosei. Lett., 162 (1993) 85-88. [4] Cdsp, T., Stafinsky, J.L., Hoskins, D.L., Perni, V.C., Uram, M. and Gordon, T.L., Age-Jelated changes in the spinal antinocieep-
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