*Corresponding author:Doctor Giuseppe Gagliardi, Department of Anesthesiology and Intensive Care, Rovigo Hospital, Italy, Email: [email protected]
How to cite this article: Gagliardi V, Gagliardi G, Lovato A, Ceccherelli F. Systemic Morphine but Not Local Administration Shows Preventive Anti-Inflammatory Effect in a Rat Model of Capsaicin-Induced Paw Edema. J Complement Med Alt Healthcare. 2023; 12(3): 555840. DOI: 10.19080/JCMAH.2023.12.555840
Peripheral opioids and analogues have demonstrated their efficacy in controlling inflammatory response and hyperalgesia. However, it is not yet clear whether the effect on edematogenic response is determined by the action at peripheral level or by systemic opioid receptors.
Methods: In this study we investigated the effects of subcutaneous, intraplantar and intraperitoneal morphine injection in rats with acute capsaicin-induced hind paw inflammation. The experiment was conducted on 50 male Sprague-Dawley rats randomly divided into 5 groups of equal numerosity. Group 1 was control group; group 2 was treated with 5 mg/kg morphine by intraperitoneal injection; group 3 with 5 mg/kg morphine injected in the back subcutaneously; group 4 and group 5 were treated respectively with 0.05 mg/kg and 0.1 mg/kg morphine locally injected in the paw before induction of neurogenic edema. Neurogenic edema was induced by the administration of 50 mcg capsaicin in 50 mcl saline in the paw. The edema was measured using a calibrated plethysmometer.
Results: Intraperitoneal injection of 5 mg/kg morphine 30 minutes before administration of capsaicin efficaciously inhibited the edematogenic response and statistically significant different variations in paw volume to the control group were measured at all observation times. The same doses of subcutaneous morphine reduced edematogenic response as well, even though to a lesser extent. Intraplantar morphine (0.05 mg/kg or 0.1 mg/kg) did not seem to have any effect on the edematogenic response at either dose or no significant difference was observed with respect to the control group.
Conclusions: the action of morphine on capsaicin-induced neurogenic edema appears to be related to a central rather than a local control mechanism.
Opioid peptides have demonstrated their efficacy in controlling inflammatory response. They exert their therapeutic action by binding to receptor sites in the central nervous system and to peripheral opioid receptors . In the periphery the opioid receptors are located on sensory fibres and post-gangliar sympathetic endings , and on other non-nervous structures, such as vascular endothelium [4,19], immunocompetent cells as lymphocytes and macrophages , and keratinocytes . The efficacy of endogenous opioids in reducing hyperalgesia and inflammatory response is extensively described in the literature . Their anti-hyperalgesic and anti-inflammatory effect in experimental rat models [22, 28] has also been demonstrated by local administration of mu agonists, morphine, and fentanyl. Similar results have been observed with the intraplantar injection of tramadol, whose analgesic potency has been shown to be greater and more durable than treatment with lidocaine . These effects are linked to tolerance and anti-hyperalgesic, anti-allodynic and anti-exudative actions are reduced when administration is
prolonged . Morphine’s site of action for edematogenic response
is not well defined. According to some authors, morphine reduces
neurogenic plasma extravasation induced by carrageenan with a
mechanism, which does not seem to be dose-dependent, and this
action is mediated primarily by peripheral opioid receptors .
Other authors report that activation of brain receptors inhibits
carrageenan-induced edema, suggesting that the anti-edematous
effect of morphine is due to action at central receptors alone
(12). Sacerdote et al.  assessed the effects of a single dose of
morphine (10μg) administered into the inflamed paw in a yeast
model of acute inflammation and observed no effect on paw edema.
Perrot et al.  administered morphine locally (50-200μg), both
before and after intraplantar carrageenan, concluding that neither
preventive nor curative administration of morphine could affect
The aim of this study is to test whether the preventive action of
morphine on edematogenic response during acute inflammation
capsaicin-induced is governed primarily by a systemic mechanism.
In this study we investigated the effects of morphine after
subcutaneous, intraplanar and intraperitoneal administration
in rats with acute capsaicin-induced hind paw inflammation.
The experiment was conducted on 50 male Sprague-Dawley
rats weighting 100-140 g (Morini Laboratories, Italy), randomly
divided into 5 groups of ten animals each. Group 1 was the
control group; group 2 was treated with 5 mg/kg morphine by
intraperitoneal administration; group 3 was treated with 5 mg/kg
morphine injected subcutaneously in the back; group 4 received
0.05 mg/kg local morphine in the paw before induction of
neurogenic edema and group 5 was treated with 0.1 mg/kg local
morphine in the paw.
The study obtained the consent of the Committee for
experimentation and was carried out at the Department of
Experimental Surgery of the University of Padua. After mild
anesthesia with ether, all the animals were given an initial
intraperitoneal injection of 400 mg/kg of chloral hydrate, and,
if necessary, received a supplemental dose of 30 mg of chloral
hydrate. After anesthesia we shaved the right hind paw of all the
animals using a hypoallergenic, non-irritant depilatory cream and
made an indelible reference mark for the volume measurements.
We then made the baseline measurement of paw volume. We
treated all groups, except the control group, with morphine at the
dose and by the administration route indicated in the protocol.
Thirty minutes later we induced neurogenic edema by injecting
50 mcg of capsaicin in 50 mcl saline in the paw. The control group
rats remained untouched for 30 minutes before the induction of
Measuring Paw Volume
We measured the edema with a calibrated plethysmometer (U.
Basile, Mod 7150) , consisting in two interconnected Perspex
cells filled with a specially prepared solution. This solution is
composed of bidistilled water, 3g/l sodium chloride, and 5ml
of a surface-active agent to reduce surface tension. The rat paw
under study is dipped in one cell while the other cell, containing
an electrode, detects variations in volume. A digital transducer
indicates small variations in volume on a luminous display. We
made three measurements for each recording and calculated the
mean of the three values. The measurements were made before
the start of treatment and every 15 minutes after induction of
edema with capsaicin for two hours.
The percentage variation with respect to baseline
measurements at the various times was calculated for each group,
according to the following formula:
%V = (Vbaset + n −Vbaset) /Vbaset ×100
The mean and standard deviation of the data collected for
various treatment groups were compared with the controls.
The statistical significance of the differences was calculated by
variance analysis (ANOVA) and t-test.
The treatment of animals conformed to the guidelines
contained in the “European Convention for The Protection of
Vertebrate Animals Used for Experimental and Other Scientific
Purposes” published on March 18, 1986, and successively modified
by the European Commission and published in the European
Official Gazzette, June 18, 2007.
The protocol was approved by the Italian Ministry of Health.
Table 1 summarizes the results, given as mean and standard
deviation (SD) of the percentage of variation of paw volume. As
shown, the edematogenic response observed after administration
of capsaicin in the control group reached its highest level after
approximately 60 minutes, remaining stable for the entire
duration of the experiment. Pre-treatment with systemic
morphine inhibited the edematogenic response. Intraperitoneal
administration of morphine (5 mg/kg) thirty minutes before
inducing edema demonstrated its efficacy by inhibiting the
edematogenic response: statistically significant differences
between this group and control group were recorded at all
observation times (Figure1).
The edematogenic response to capsaicin was reduced also
in the group, which received the same dose of morphine (5 mg/kg) subcutaneously: also, this group showed a statistically significant difference (p<0.05) compared to the control group at
all observation times, though the effect was less marked than the
group treated with intraperitoneal injection (Figure 1).
Intraplantar morphine did not seem to have any effect on
edematogenic response at either administered dose (0.05 mg/
kg and 0.1 mg/kg), as no statistically significant difference could
be detected when these groups were compared to control group
(Figure 2). On the contrary a statistically significant difference was
present when these groups were compared to rats treated either
with intraperitoneal or subcutaneous injection (Figure 3).
The results of our study seem to confirm that systemic
morphine has a preventive effect on capsaicin induced edema. In
fact, all the rats receiving 5 mg/kg morphine showed a reduced
edematogenic response to capsaicin injection if compared to
control animals. Intraperitoneal route seemed to be more effective
than subcutaneous injection, maybe for a lower bioavailability of
the latter due to slower absorption.s
Local intraplantar injection of morphine (0.05 mg/kg or 1
mg/kg) did show no preventive effect on edematogenic response to capsaicin. The effectiveness of local opioid administration in controlling pain of neurogenic inflammation is well documented
and is due to opioid binding to receptors present on nervous
endings of the inflamed part .
In the experimental model of the present work edema is due to
the direct action of the irritant on the capsaicin-sensitive nervous
fibers, which are small peptidergic neurons (50 μm) functionally
corresponding to unmyelinated (C) fibers . Capsaicin binds to
the transient receptor potential vanilloid-1 (TRPV1) on primary
nociceptive fibers and stimulates the release of substance P and
calcitonin gene-related peptide  by a centrally mediated
antidromic activity of Aδ and C primary afferent fibers [15, 16].
Dorsal root reflexes (DRRs) play a foundamental role for
antidromic activity of small sensitive fibers during neurogenic
inflammation induced by capsaicin. Capsaicin also exerts a direct
action on small fibers: it elicits axo-axonic reflexes possibly
contributing to inflammation at the site of injection . So,
peripheral administration of opioids can be effective in controlling
pain but inadequate to prevent the reflex edematogenic response.
As reported by Obara, in an experimental model of carrageenaninduced
inflammation, the use of peripherically acting opioids,
intraplantar morphine or 6-amino acid-substituted derivatives,
elicits a potent, long-lasting analgesic response, but does not show
any significant influence on the extent of the edematogenic effect
. Our results seem to confirm these data as local injection
of morphine did not have any influence on the edematogenic
response to capsaicin. Otherwise, the acute systemic
administration of morphine can act on the tissue inflammatory
response to surgical incision by modulating neutrophiles
infiltration and cytokines accumulation in the tissue surrounding
the surgical wound . These results point to the foundamental role of central mechanisms in eliciting and maintaining neurogenic inflammation. The concept of preventive analgesia is based on
the observation that an intense stimulation of the nociceptive
primary afferent fibers, due to tissue damage and inflammation,
increases the eccitability of the nociceptive spinal dorsal horn
neurons so contributing to post-injury pain and hyperalgesia [3,6]
and . An early preventive action on the nociceptive inputs
to the spinal cord decreases hypereccitability, hence pain and
hyperalgesia. Therefore, the preventive use of systemic morphine
exerts its analgesic effect and, by acting on neurons of the dorsal
horns, modulates neurogenic inflammation triggered by capsaicin
sensitive fibers through reflexes at this anatomical level .
The clinical implication of this observation is that preventive
morphine may prove useful in the treatment of pain associated
with inflammation, such as after surgery, in which the combination
of these effects probably takes part to the observed improved