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Vol. 296, Issue 2, 270-275, February 2001
Department of Applied Pharmacology, Faculty of Pharmaceutical Sciences (I.T., H.N., Y.K.) and Department of Virology, Faculty of Medicine (K.S.), Toyama Medical and Pharmaceutical University, Toyama, Japan; and National Heart, Lung, and Blood Institute, National Institute of Health, Bethesda, Maryland (T.A.)
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Abstract |
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 Top
 Abstract
 Introduction
Materials and Methods
Results
Discussion
References
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The effects of systemic and local injections of gabapentin, a novel anticonvulsant agent, were tested on nociceptive behaviors in mice with acute herpetic pain. Transdermal infection with herpes simplex virus type-1 (HSV-1) produced nociceptive hypersensitivity of the infected hind paw to innocuous (allodynia) and noxious mechanical stimulation (hyperalgesia) with von Frey filaments. Systemic administration of gabapentin (10-100 mg/kg, peroral) produced a dose-dependent inhibition of both allodynia and hyperalgesia; gabapentin (30-300 mg/kg) did not affect locomotor activity. Intrathecal injection of gabapentin (10-100 µg/animal) also attenuated dose dependently both nociceptive hypersensitivities. In contrast, intraplantar, intracisternal, and intracerebroventricular administration of gabapentin (10-100 µg/animal) had no effect on the HSV-1-induced nociceptive hypersensitivities. Pretreatment with naltrexone (1 mg/kg) inhibited antinociceptive effect of morphine (5 mg/kg), but not gabapentin (100 mg/kg). Repeated administration of morphine (5 mg/kg, four times) led to tolerance of antinociceptive action, whereas gabapentin (100 mg/kg, four times) had antinociceptive effect even after the forth administration. The present results suggest that gabapentin is effective in the treatment of acute herpetic pain without apparent adverse effects, and analgesic action of gabapentin is mainly mediated by actions on the spinal cord.
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Introduction |
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Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
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Gabapentin
[1-(aminomethyl)cyclohexaneacetic acid], a novel anticonvulsant
agent, is currently in clinical use as an "add-on" therapy in
patients with partial seizures resistant to conventional therapies (Goa
and Sorkin, 1993
). Although gabapentin is a synthetic analog of
-aminobutyric acid (GABA) and easily penetrates into the central
nervous system, it does not interact with either
GABAA or GABAB receptors
(Taylor et al., 1998). It has been recently reported that gabapentin
alleviates neuropathic pain, including postherpetic neuralgia, reflex
sympathetic dystrophy, and diabetic neuropathy (Mellick et al., 1995;
Rosner et al., 1996; Rosenberg et al., 1997). The clinical efficacy has
been supported by studies using various animal models of pain. For
example, systemic injection of gabapentin reverses allodynia of rats
with neuropathy induced by partial ligation of the sciatic nerve (Pan
et al., 1999) and prevents the development of thermal hyperalgesia in a
rat model of postoperative pain (Field et al., 1997). Moreover, local
(spinal or peripheral) injection of gabapentin reduces formalin-induced nociceptive behaviors (Carlton and Zhou, 1998; Yoon and Yaksh, 1999).
Thus, there are several lines of evidence for effectiveness of
gabapentin in the management of various pain in animals and human
subjects; however, the analgesic mechanisms and site of action of
gabapentin are still unknown.
Recently, we have developed a mouse model of acute herpetic pain
(Takasaki et al., 2000a). When inoculated with herpes simplex virus
type-1 (HSV-1) on the skin of the hind paw of the mouse, zosteriform
skin lesions developed unilaterally in a corresponding dermatome after
a 4-day latent period. Such mice showed aversive responses to innocuous
tactile stimulation (designated as allodynia) and noxious mechanical
stimulation (designated as hyperalgesia). These nociceptive responses
became apparent when HSV-1 replicated in the dorsal root ganglion and
eruption developed on the skin. We showed that HSV-1-induced allodynia
and hyperalgesia were attenuated dose dependently by systemic (peroral)
administration of gabapentin (Takasaki et al., 2000b). Thus, this study
was conducted to determine the site and characteristics of
antinociceptive action of gabapentin in mice with acute herpetic pain.
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Materials and Methods |
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Abstract
Introduction
Materials and Methods
Results
Discussion
References
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Animals. Female BALB/c mice weighing about 20 g (6 weeks old at the start of experiments; Japan SLC, Shizuoka, Japan) were used. They were housed six per cage under controlled temperature (22 ± 1°C) and humidity (55 ± 10%). The room was lighted from 7:00 AM to 7:00 PM and during the behavioral test. Food and water were freely available. HSV-1 inoculation and behavioral experiments were done in the infection room of Molecular Genetics Research Center, Toyama Medical and Pharmaceutical University, Toyama, Japan.
Virus Infection.
The mice were inoculated with HSV-1 as
described (Takasaki et al., 2000a). Briefly, HSV-1 (7401H strain,
1 × 106 plaque-forming units in 10 µl)
was inoculated on the depilated shin skin of the right hind paw (5 × 5 mm) after scarification with 27-gauge needles. The contralateral
hind paw was without inoculation.
Behavioral Test.
Allodynia and hyperalgesia of the hind paw
were assessed as described (Takasaki et al., 2000a). The mice were
placed individually in a plastic cage (11 × 18 × 15 cm)
with a wire mesh bottom. After acclimation period of at least 15 min,
von Frey filaments with bending forces of 0.03 g (innocuous
stimulation) and 1.20 g (noxious stimulation) were pressed
perpendicularly against the plantar skin and held for 3 to 5 s
with it slightly buckled. The stimulation of the same intensity was
applied six times to each hind paw at intervals of several seconds. The
responses to these stimuli were ranked as follows: 0, no response; 1, move away from von Frey filament; and 2, immediate flinching or licking
of the hind paw. Nociceptive score was calculated as follows:
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).
Drug Administration.
Gabapentin was synthesized at
Department of Organochemical Design and Synthesis, Faculty of
Pharmaceutical Sciences, Toyama Medical and Pharmaceutical University
(Takasaki et al., 2000b). It was dissolved in tap water for peroral
injection or in physiological saline for intrathecal (Hylden and
Wilcox, 1980), intraplantar, intracisternal (Ueda et al., 1979), and
intracerebroventricular (Haley and McCormick, 1957) injections.
Intrathecal, intracisternal, and intracerebroventricular injections
were given in a volume of 5 µl and intraplantar injection was given
in a volume of 20 µl. Morphine hydrochloride (Sankyo, Tokyo, Japan)
was dissolved in physiological saline and administered s.c. Effects of
gabapentin and morphine were tested on day 6 and in some case on day 7 postinoculation. The µ-opioid receptor antagonist naltrexone (Sigma,
St. Louis, MO) was dissolved in physiological saline and administered
s.c. at a dose of 1 mg/kg 15 min before morphine and gabapentin injection.
Locomotor Activity Test. Mice were placed individually in a wheel cage (25 cm in diameter and 6 cm in width), and locomotor activity was measured as the number of wheel revolutions. Locomotor activity was assessed from 1 to 2 h after gabapentin administration.
Data Analysis. Unless otherwise mentioned, the means of data are presented together with S.E.M. Results of behavioral experiments were analyzed with one-way ANOVA or repeated measures two-way analysis of variance and post hoc Dunnett's multiple comparisons; p < 0.05 was considered significant.
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Results |
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Abstract
Introduction
Materials and Methods
Results
Discussion
References
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HSV-1 inoculation on the unilateral hind paw of mouse produced
allodynia (nociceptive response to 0.03-g von Frey filament) and
hyperalgesia (nociceptive response to 1.20-g von Frey filament). Allodynia and hyperalgesia of the HSV-1-inoculated hind paw became apparent since day 5 postinoculation and persisted at least until day 8 (Fig. 1). HSV-1 inoculation did not
affect the responses of the contralateral (uninoculated) hind paw to
von Frey filaments until at least day 8 postinoculation (Fig. 1).
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Effects of Systemic Gabapentin on HSV-1-Induced Allodynia and
Hyperalgesia.
Systemic administration of gabapentin (10-100 mg/kg
p.o.) produced a significant, dose-dependent inhibition of allodynia
and hyperalgesia induced by HSV-1 infection; the 100-mg/kg dose of gabapentin produced complete and partial inhibition in allodynia and
hyperalgesia, respectively (Fig. 2, A and
B). In contrast, gabapentin did not affect the behavioral responses of
the contralateral (uninfected) hind paw to von Frey filaments (Fig. 2,
C and D). Locomotor activity was not affected by antinociceptive and
higher doses of gabapentin (30-300 mg/kg) (Fig.
3).
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Site of Antinociceptive Action of Gabapentin.
Intrathecal
injection of gabapentin (10-100 µg/animal) produced a significant,
dose-dependent inhibition of both allodynia and hyperalgesia (Fig.
4, A and B). The inhibitory effects of the highest dose (100 µg/animal) peaked 60 min after injection and
subsided by 120 min. Intrathecal gabapentin had no effect on the
response of the contralateral hind paw at doses of 10 to 100 µg/animal (data not shown). Intraplantar, intracisternal, and
intracerebroventricular injections of gabapentin did not affect allodynia and hyperalgesia induced by HSV-1 infection (Fig.
5, C-H). Any abnormal behaviors,
including decrease or increase of locomotor activity, were not observed
after local injections of gabapentin.
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Effect of Naltrexone on Antinociceptive Action of Gabapentin and
Morphine.
The inhibitory effects of gabapentin (100 mg/kg p.o.) on
allodynia and hyperalgesia induced by HSV-1 infection were not affected by pretreatment with naltrexone (1 mg/kg s.c., 
15 min) (Fig. 6, A and B). Morphine (5 mg/kg s.c.)
attenuated both allodynia and hyperalgesia induced by HSV-1 infection.
The effects of morphine were almost completely antagonized by
pretreatment with naltrexone (1 mg/kg s.c.) (Fig. 6, C and D).
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Tolerance to Antinociceptive Effects.
Repeated administration
of gabapentin (100 mg/kg p.o., twice a day) produced constant
inhibition on allodynia and hyperalgesia for at least 2 days; the
antinociception was apparent even after the fourth administration (Fig.
7, A and B). On the other hand, the
antinociceptive effects of morphine (5 mg/kg s.c.) were rapidly decreased after repeated administration (twice a day); the effects of
the third and fourth administration were significantly smaller than
that of the first administration (Fig. 7, C and D).
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Discussion |
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Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
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The mice infected with HSV-1 exhibited allodynia and hyperalgesia
since day 5 postinoculation and such nociceptive hypersensitivities persisted until at least day 8, confirming our previous observation (Takasaki et al., 2000a). The occurrence of the nociceptive
hypersensitivity may be due to the infection and especially
proliferation of HSV-1 in the dorsal root ganglia, because the amount
of HSV-1 DNA (Takasaki et al., 2000a) and the number of HSV-1
antigen-positive cells (Takasaki et al., 2000b) in the dorsal root
ganglia peaked on day 5 postinoculation.
In the present study, systemic injection of gabapentin markedly
attenuated both allodynia and hyperalgesia induced by HSV-1 infection.
Peroral administration of antinociceptive doses (30 and 100 mg/kg) and
even the higher dose (300 mg/kg) of gabapentin did not affect locomotor
activity, suggesting that gabapentin does not have sedative and motor
deficit effect at doses tested. Thus, antiallodynic and
antihyperalgesic actions of gabapentin may not be mediated by sedation
or motor deficits. Importantly, in patients with postherpetic neuralgia
(Segal and Rordorf, 1996), idiopathic trigeminal neuralgia (Sist et
al., 1997) or painful diabetic neuropathy (Gorson et al., 1999),
gabapentin eliminates pain without serious side effects such as
somnolence or dizziness that cause either discontinuation or reduction
of the dose.
One important finding in this study is that only intrathecal, but not
intraplantar, intracisternal, and intracerebroventricular injection of
gabapentin was effective against tactile-evoked allodynia and
mechanical hyperalgesia in mice infected with HSV-1, suggesting that
the antiallodynic and antihyperalgesic actions of gabapentin are
mediated primarily by the action on the spinal cord, but not on
peripheral and supraspinal regions. Coincident with our results, intrathecal gabapentin is effective on tactile-evoked allodynia in rats
with surgically induced neuropathic pain (Hwang and Yaksh, 1997) and on
formalin-induced pain behavior in the rat (Yoon and Yaksh, 1999).
Because high levels of specific binding of
[3H]gabapentin were detected in the outer
layers of the cerebral cortex (Hill et al., 1993), the brain is
considered to be a site of antinociceptive action. In the present
study, however, intracisternal and intracerebroventricular injections
of gabapentin failed to attenuate both allodynia and hyperalgesia,
suggesting that the brain is not a primary site of antinociceptive
action of gabapentin at least in acute herpetic pain.
It has been reported that intraplantar injection of gabapentin (6-600
µg/animal) attenuates nociceptive behavior induced by formalin
injection (Carlton and Zhou, 1998), suggesting the peripheral action of
gabapentin. In this study, however, local injection of gabapentin into
the region of stimulation had no effect on acute herpetic pain. In our
mouse model of herpetic pain, nociceptive response may be mainly due to
the propagation of HSV-1 in the dorsal root ganglia, but not to the
peripheral inflammation (Takasaki et al., 2000a). Peripheral injection
of gabapentin may be effective against the pain of peripheral
inflammation, but not to neuropathic pain.
In contrast to morphine, antinociceptive action of gabapentin was not
blocked by pretreatment with naltrexone, a µ-opioid receptor
antagonist, suggesting that antiallodynic and antihyperalgesic effects
of gabapentin are not mediated by µ-opioid receptors. Furthermore,
although repeated administration of morphine produced apparent
tolerance to its antinociceptive effects, tolerance did not develop to
the antinociceptive action of gabapentin. These results are consistent
with previous report that repeated administration of gabapentin for 6 days did not lead to development of tolerance to the antinociceptive
effects in formalin test (Field et al., 1997). It is noteworthy that
the duration of antinociceptive effects of gabapentin is longer than
that of morphine (Takasaki et al., 2000b).
In the present study, gabapentin did not affect the behavioral
responses of the contralateral (uninoculated) hind paw to von Frey
filaments. It was reported that gabapentin (30-300 mg/kg s.c.) failed
to show an antinociceptive effect in transient pain models in rats
(Field et al., 1997). Intrathecal gabapentin (30-300 µg/animal) also
has no effects on the thermal escape latency of the normal hind paw in
rats (Jun and Yaksh, 1998). Similarly, in clinical use, gabapentin does
not inhibit pain induced by strong stimuli, although it was claimed to
be particularly effective against tactile- and cold-allodynia (Attal et
al., 1998). Therefore, gabapentin seems to have antiallodynic and
antihyperalgesic effects but not antinociceptive effect on
nociceptor-specific pain; gabapentin may be effective against
nociceptive hypersensitivity induced by tissue damage or neuropathy.
There are many pharmacological investigations about gabapentin;
however, the mechanisms of antiallodynic and antihyperalgesic action of
gabapentin are still unknown. It has been shown that gabapentin binds
with a high affinity to the 
2
-subunit of
voltage-dependent calcium channel (Gee et al., 1996). Although the
physiological role of 2-subunit is not well
understood, it appears to be common to all types of voltage-dependent
calcium channels. It has been reported that intrathecal application of
N-type voltage-dependent calcium channel antagonist attenuates
mechanical allodynia induced by tight ligation of L5 and L6 spinal
nerves in rats (Chaplan et al., 1994), and that N- and P-type channel
antagonists produced antinociception in formalin test in rats (Malmberg
and Yaksh, 1994; Diaz and Dickenson, 1997). These findings raise the
possibility that the subunit of the voltage-dependent calcium channel
may play an important role in mechanism of action of gabapentin.
Another potential of analgesic mechanisms of gabapentin is the blockade
of glutamatergic systems and NMDA receptor in spinal dorsal horn.
Recently, it has been reported that gabapentin decreases the amplitudes
of both non-NMDA and NMDA receptor-mediated excitatory postsynaptic
currents in dorsal horn neurons of the rat spinal cord slice (Shimoyama
et al., 2000). Furthermore, it has been reported that
D-serine, an antagonist at the strychnine-insensitive glycine site on the NMDA receptor, inhibits antihyperalgesic effects of
gabapentin in thermal injury rats (Jun and Yaksh, 1998). These findings
suggest that gabapentin may also have an inhibitory effect on
glutamatergic excitatory neurotransmission in the dorsal horn. Voltage-dependent calcium channels (N- and P-types) localized at
synaptic sites are involved in the release of transmitters, such as
glutamate (Turner et al., 1993). Thus, blockade of voltage-dependent calcium channels and inhibition of glutamate release in the spinal dorsal horn may be involved in the antinociceptive action of
gabapentin. Whether glutamatergic release is increased in the spinal
dorsal horn of HSV-1-infected mice and whether gabapentin attenuates glutamate release are interesting questions to be elucidated in future studies.
In summary, we demonstrated that gabapentin markedly attenuated nociceptive hypersensitivity induced by HSV-1 infection in mice; the effect may be mediated primarily by the spinal action. Tolerance did not develop to the antinociceptive action of gabapentin and behavioral abnormalities such as sedation and motor dysfunction were not apparent after antinociceptive doses of gabapentin. Thus, gabapentin may be useful in the treatment of the acute herpetic pain.
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Acknowledgments |
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We thank Drs. Hideo Nemoto and Hiroki Takahata for the synthesis of gabapentin.
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Footnotes |
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Accepted for publication October 17, 2000.
Received for publication August 16, 2000.
Send reprint requests to: Yasushi Kuraishi, Ph.D., Department of Applied Pharmacology, Faculty of Pharmaceutical Sciences, Toyama Medical and Pharmaceutical University, 2630 Sugitani, Toyama 930-0194, Japan. E-mail: kuraisiy{at}ms.toyama-mpu.ac.jp
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Abbreviations |
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GABA, -aminobutyric acid;
HSV-1, herpes
simplex virus type-1;
NMDA, N-methyl-D-aspartate.
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References |
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Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
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2 subunit of a calcium channel.
J Biol Chem
271:
5768-5776-aminobutyric acid on the hyperalgesia observed after thermal injury in the rat.
Anesth Analg
86:
348-354[Abstract].This article has been cited by other articles:
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  C. Hansen, I. Gilron, and M. Hong The Effects of Intrathecal Gabapentin on Spinal Morphine Tolerance in the Rat Tail-Flick and Paw Pressure Tests Anesth. Analg., October 1, 2004; 99(4): 1180 - 1184. [Abstract] [Full Text] [PDF]
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 I. Takasaki, T. Suzuki, A. Sasaki, K. Nakao, M. Hirakata, K. Okano, T. Tanaka, H. Nagase, K. Shiraki, H. Nojima, et al. Suppression of Acute Herpetic Pain-Related Responses by the {kappa}-Opioid Receptor Agonist (-)-17-Cyclopropylmethyl-3,14{beta}-dihydroxy-4,5{alpha}-epoxy-6{beta}-[N-methyl-3-trans-3-(3-furyl) Acrylamido] Morphinan Hydrochloride (TRK-820) in Mice J. Pharmacol. Exp. Ther., April 1, 2004; 309(1): 36 - 41. [Abstract] [Full Text]
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), 30 (
), and 100 µg (
) and
vehicle (saline; 
) were injected intrathecally on day 6 postinoculation. The data presented are means and S.E.M.
(n = 6). Repeated measures-ANOVA analysis: main
effect of gabapentin, F(3,8) = 5.41, p < 0.01; interaction between gabapentin and time,
F(24,160) = 3.44, p < 0.001 (A); and main effect of gabapentin, F(3,8) = 7.51, p < 0.01; interaction between gabapentin and time,
F(24,160) = 6.59, p < 0.0001 (B).
). Effect of gabapentin was
assessed 30 min after injection. The data presented are means and
S.E.M. (n = 6). *p < 0.05 compared with S (Dunnett's test).
