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Antinociceptive effect of mirtazapine in rats with diabetic neuropathy.


Diabetic neuropathy is a chronic complication associated with diabetes that has a significant detrimental effect on the daily activities of patients. It occurs in approximately 50% of patients with diabetes within 20 years of diagnosis (1). Different drugs have been used in the treatment of neuropathy such as tricyclic antidepressants (amitriptyline), selective serotonin reuptake inhibitors (fluoxetine), antiepileptics (carbamazepine, phenytoin), venlafaxine, amantadine, tramadol, oxycodone, gabapentin, bupropion, and capsaicin (1). Mirtazapine is a new antidepressant that acts in a manner that is different from other antidepressants and is notable for the low frequency of severe side effects when compared with other antidepressants (2).

Mirtazapine blocks the presynaptic [alpha]-2 adrenoreceptors in both the central and peripheral nervous systems, while its affinity to [alpha]-1 adrenoreceptors is less marked. In addition, it weakly blocks serotonin-1 (5-hydroxytryptamine-1,5-HT-1) receptors and strongly blocks 5-HT-2 and 5-HT-3 receptors. On the other hand, it has no effect on noradrenaline re-uptake and does not block the subtypes of [beta]-adrenergic receptors to any significant degree (2).

Mirtazapine, with its low side-effect profile, might be considered a good alternative in the treatment of diabetic neuropathy. This study investigated the antinociceptive effect of mirtazapine on diabetic neuropathy in rats and the role of opioidergic, serotoninergic, and adrenergic systems in this effect. Naloxone, a competitive opioid [mu] (mu) receptor antagonist that removes the respiratory depression effect resulting from opioid toxicity, was used to evaluate the role of the opioidergic system (3). Furthermore, a non-selective serotonin receptor antagonist, metergoline (4), and a [alpha]-2a adrenergic receptor antagonist, BRL44408 (5), were used to evaluate the roles of the serotoninergic and adrenergic systems, respectively


This study was approved by the Local Ethics Board of Animal Research. Sprague Dawley rats weighing 170-225 g were used in the study They were divided into II groups, each containing four males and four females. Rats were kept in a 12-h daylight and 12-h darkness cycle at a room temperature of 20[degrees]C and a humidity of 50%-60% and were administered standard pellet feed and water Rats were brought to the experiment area I h prior to the start of experiments to enable them to adjust to the environment.

Sensorimotor Performance

A rotarod test was used to detect the motor activity of rats to be included in the study, with the adequacy of motor activity being defined as being able to stay on the rod for at least 120 s.

Nociceptive Test (Hot-plate Test)

Rats were placed on a hot plate after the plate temperature had been brought to 52.5[degrees]C. The length of time from the placement of the rat on the plate until it shook or rapidly withdrew its posterior leg was measured using a chronometer

Drugs and Chemical Substances

Streptozotocin (STZ; Sigma-Aldrich, St. Louis, MO, USA) and naloxone (Sigma-Aldrich, St. Louis, MO, USA) were dissolved in physiological saline; mirtazapine (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in 10% polyethylene glycol; and metergo-line (Sigma-Aldrich, St. Louis, MO, USA) and BRL44408 (Sigma-Aldrich, St. Louis, MO, USA) were dissolved in 10% ethanol. The solutions were immediately prepared prior to administration and were intraperitoneally injected in volumes of 0.3-0.45 mL.

Experimental Procedures

Basal hot-plate test measurements were conducted during the initial stage of the experiment. Mirtazapine was administered before the development of diabetes at doses of 5, 10, and 15 mg/kg. Subsequently STZ was intraperitoneally injected at a dose of 50 mg/kg to enable the development of diabetes (6,7). The glucose level in the tail vein blood was measured on the third and fifth days using a glucometer (Accu-Chek Active). Rats with a blood glucose level of 240 mg/dL or higher were considered as being diabetic.

After a 3-week waiting period, the second hot-plate test measurements were conducted to check whether hyperalgesia secondary to diabetic neuropathy had developed. The development of hyperalgesia was diagnosed in rats as a 20% decrease in measurements compared with the basal values. Drugs specific to the experimental groups were administered to rats with hyperalgesia (Table 1). Furthermore, hot-plate tests were performed at 30 and 60 min after drug administration.

Statistical Analysis

The Shapiro-Wilk test was used to evaluate the normal distribution. Because the data were normally distributed, the differences between the groups were evaluated using a variance analysis. Dunnet's test was used to compare the nociceptive measurement values of the control group and the other 10 groups. A paired t-test was used for the paired comparison of the in-group values that were found to be significant, with a p value of <0.05 being accepted as significant.


Effects because of STZ Application

Approximately 75% of rats that were administered STZ developed hyperglycemia (diabetes). The body weights of rats with diabetes were significantly reduced compared with their pre-diabetic status (p<0.05; Table 2). The hot-plate test values of the group that was administered only STZ were not significantly different from those of the control group (Figure 1). The hot-plate test values obtained 3 weeks after STZ administration were decreased by approximately 20% when compared with the pre-injection values. This decrease was accepted as a sign of the development of hyperalgesia in rats (p<0.05; Table 3).

Effects of Mirtazapine and Antagonists

The tests performed on the rotarod equipment demonstrated that the drugs caused no impairment in locomotor activity (Table 4). Mirtazapine (10 and 15 mg/kg) was observed to have a significant antinociceptive effect in rats that had not yet developed diabetes (p<0.05; Figure 2). Mirtazapine (15 mg/kg) was also found to have a significant antinociceptive effect in rats that had developed hyperalgesia because of diabetic neuropathy (p<0.05; Figure 3). Naloxone, metergoline, and BRL44408 inhibited the antinociceptive effect of mirtazapine to a significant degree (15 mg/kg); however the combination of mirtazapine and other drugs still demonstrated an antinociceptive effect when compared with the control group (p<0.05; Figure 4). These drugs were observed to have no different effects when applied as a single agent when compared with the control group (Figure 5). Furthermore, polyethylene glycol and ethanol, which were used to dissolve metergoline and BRL44408, respectively did not produce any different effects when used as a single agent (Figure 6).


The antinociceptive effect of mirtazapine in rats with STZ-induced diabetic neuropathy was investigated in this study. Although the diabetes-inducing rate of STZ has not been cited in the literature, cases with no induction of diabetes have been reported in almost all studies (8,9).

Diabetic neuropathy is one of the most common complications associated with diabetes and can be detrimental to the quality of life of the patient. Thermal, chemical, and mechanical hyperalgesia have all been reported in patients who have developed diabetic neuropathy (10). There are different opinions in the literature regarding the development of thermal hyperalgesia in experimental diabetes induced with STZ; some researchers claim that there is no change in thermal sensitivity in experimental diabetes, while others have reported the development of thermal hyperalgesia (10,11). In this study, thermal hyperalgesia, which was demonstrated by the hot-plate test, was accepted as a sign of diabetic neuropathy In previous studies, it has been demonstrated that experimental diabetic neuropathy occurs in 2-3 weeks following diabetes induction by STZ (12); the development of diabetic neuropathy was also detected within approximately 3 weeks in this study. Pain measurement is achieved through nociceptive stimulus in animal experiments and may be thermal, mechanical, or chemical in nature. The antinociceptive effects of some antidepressant drugs were evaluated using different pain models in a study by Bomholt et al. (13), who found that amitriptyline, duloxetine, mirtazapine, and citalopram were found to be ineffective in a tail withdrawal test, which is an acute pain model; in contrast, duloxetine and mirtazapine were demonstrated to have a significant antinociceptive effect in a hot-plate test. Therefore, the hot-plate test method was selected for this study.

Mirtazapine demonstrated to have a marked antinociceptive effect in rats in this study with a dose range of 5-15 mg/kg demonstrating dose-dependent antinociceptive efficacy. In a study of rats by Schreiber et al. (14) using a hot-plate test, mirtazapine demonstrated an antinociceptive effect at a dose of 10 mg/kg; however it was found to be partially ineffective at a dose of 15 mg/kg. This difference may be attributed to the type of animals used in the studies.

The antinociceptive effect that was induced by mirtazapine was partially antagonized by naloxone [opioidergic system contribution; (14,15)], metergoline (serotoninergic system contribution; 14), and BRL44408 [adrenergic system contribution; (14,16)] in this study. Compatible with our results, Schreiber et al. (14) suggested that opioidergic, serotoninergic, and adrenergic systems mediated the antinociceptive effect of mirtazapine. Naloxone has demonstrated antinociceptive efficacy in studies performed in rats (17,18). In this study, when administered as a single agent, naloxone had no antinociceptive effect; however it significantly decreased the antinociceptive efficacy of mirtazapine (15 mg/kg). This suggests that the analgesic effect of mirtazapine is partially mediated by the opioidergic system. There are two studies in the literature suggesting that antidepressants have a direct effect on opioid receptors and that these receptors partially mediate the analgesic effects of these antidepressants (19,20). The fact that the antinociceptive effect of clomipramine is blocked by naloxone and naltrexone, both of which are opioid receptor antagonists, was demonstrated as a proof of these suggestions (21). In many previous studies, the mechanisms of the antinociceptive effect of new-generation antidepressants have been investigated (17,18,22), and it was found that most of these drugs increased the effects of small doses (with no analgesic action) of opioids. Similarly venlafaxine, which is a (powerful) serotonin, (moderate) noradrenaline, and (weak) dopamine reuptake inhibitor mediates its antinociceptive effect through both opioidergic and adrenergic systems (23). The findings of all these studies and this study underline the role of the opioidergic system in the antinociceptive effect of mirtazapine. Mirtazapine increases serotoninergic and adrenergic neurotransmission through the blockage of serotonin receptors of the 5-HT2 and 5-HT3 type, as well as of auto and heteroadrenoreceptors (2). The net effect is the stimulation of post-synaptic 5-HT1 type serotonin receptors. In this study, metergoline demonstrated no antinociceptive effect by itself; however it was found to decrease the antinociceptive effect of mirtazapine significantly when the two were administered together Serotoninergic neurons end in the posterior horn descending from the brain stem, through the dorsolateral funiculus and up to the medulla spinalis (descending inhibitor pathway), where they play a role in the modulation of pain. This event is reversed by serotonin and adrenergic receptor antagonists (4). Metergoline may block the nociceptive effect of mirtazapine through the mechanism described above.

In this study; the role of the adrenergic system on the mechanism of the antinociceptive effect of mirtazapine was also investigated, in addition to the opioidergic and serotoninergic systems. BRL44408, an [alpha]-2 adrenergic receptor antagonist, demonstrated no antinociceptive effect by itself, but blocked significanty the antinociceptive effect of mirtazapine when the two were applied together This suggests that the adrenergic system also plays a role in the antinociceptive effect of mirtazapine. Schreiber et al. (14) also demonstrated the role of the noradrenergic system in the nociceptive effect of mirtazapine, in addition to the opioidergic and serotoninergic systems. Similarly analgesia induced by tricyclic antidepressants was reported to be removed by an [alpha]-2 adrenergic receptor antagonist, RX 821002 (24). Ghelardini et al. (25) reported that the antinociception induced by tricyclic antidepressants was inhibited by reserpine (a drug that decreases the noradrenaline release from adrenergic nerve endings) and yohimbine, an [alpha]-2 adrenergic receptor antagonist), in their study in rats using abdominal contraction and hot-plate tests. The results of all these studies and this study suggest that the adrenergic system plays a role in the antinociceptive effect of mirtazapine.

DOI: 10.5152/npa.2015.8791

Conflict of Interest: No conflict of interest was declared by the authors.

Financial Disclosure: The authors declared that this study has received no financial support.


(1.) Ziegler D, Keller J, Maier C, Pannek J. Diabetic Neuropathy. Exp Clin Endocrinol Diabetes 2014; 122:406-415. [CrossRef]

(2.) Agargun Y Ebrinc S. (Mirtazapin: A review). Klinik Psikofarmakoloji Bulteni 1998; 8:59-68.

(3.) Sabzghabaee AM, Eizadi-Mood N, Yaraghi A, Zandifar S. Naloxone therapy in opioid overdose patients: intranasal or intravenous? A randomized clinical trial. Arch Med Sci 2014; 10:309-314. [CrossRef]

(4.) Dogrul A, Seyrek M. Systemic morphine produce antinociception mediated by spinal 5-HT7, but not 5-HTIA and 5-HT2 receptors in the spinal cord. Br J Pharmacol 2006; 149:498-505. [CrossRef]

(5.) Miksa M, Das P Zhou M, Wu R, Dong W, Ji Y, Goyert SM, Ravikumar TS, Wang P Pivotal Role of the [alpha]2A-Adrenoceptor in Producing Inflammation and Organ Injury in a Rat Model of Sepsis. PLoS One 2009; 4:5504. [CrossRef]

(6.) Majithiya JB, Balaraman R, Giridhar R, Yadav MR. Effect of bis[curcumino]oxovanadium complex on non-diabetic and streptozotocin-induced diabetic rats. J Trace Elem Med Biol 2005; 18:211-217. [CrossRef]

(7.) Hwang HJ, Kim SW, Lim JM, Joo JH, Kim HO, Kim HM. Hypoglycemic effect of crude exopolysaccharides produced by a medicinal mushroom Phellinus baumii in streptozotocin-induced diabetic rats. Life Sci 2005; 76:3069-3080. [CrossRef]

(8.) Eidi M, Eidi A, Zamanizadeh H. Effect of Salvia officinalis L. leaves on serum glucose and insulin in healthy and streptozotocin-induced diabetic rats. J Ethnopharmacol 2005; 100:310-3. [CrossRef]

(9.) Distinct vasculopathic profile of major key organs. Eur J Phar 2005; 514:69-78. [CrossRef]

(10.) Lynch JJ 3rd, Jarvis ME Kowaluk EA. An adenosine kinase inhibitor attenuates tactile allodynia in a rat model of diabetic neuropathic pain. Eur J Phar 1999; 364:141-146. [CrossRef]

(11.) Malcangio M, Tomlinson DR. A pharmacologic analysis of mechanical hyperalgesia in streptozotocin/diabetic rats. Pain 1998; 76:151-157. [CrossRef]

(12.) Eox A, Eastwood C, Gentry C, Manning D, Urban L. Critical evaluation of the streptozotocin model of painful diabetic neuropathy in the rat. Pain 1999; 81:307-316. [CrossRef]

(13.) Bomholt SE Mikkelsen JD, Blackburn-Munro G. Antinociceptive effects of the antidepressants amitriptyline, duloxetine, mirtazapine and citalopram in animal models of acute, persistent and neuropathic pain. Neuropharmacology 2005; 48:252-263. [CrossRef]

(14.) Schreiber S, Rigai T Katz Y Pick CG. The antinociceptive effect of mirtazapine in mice is mediated through serotonergic, noradrenergic and opioid mechanisms. Brain Res Bull 2002; 58:601-605. [CrossRef]

(15.) Wang JW, Lundeberg T Yu LC. Antinociceptive role of oxytocin in the nucleus raphe magnus of rats, an involvement of [beta]-opioid receptor Regul Pept 2003; 115:153-159. [CrossRef]

(16.) Khasar S, Green P Chou B, Levine JD. Peripheral nociceptive effects of alfa2-adrenergic receptor agonists in the rat. Neuroscience 1995; 66: 427-432. [CrossRef]

(17.) Capuano A, De Corato A, Treglia M, Tringali G, Curro D, Dello Russo C, Navarra P Peripheral antinociceptive effects of low doses of naloxone in an in vivo and in vitro model of trigeminal nociception. Neuropharmacology 2010; 58:784-792. [CrossRef]

(18.) Wheeler-Aceto H, Cowan A. Naloxone causes apparent antinociception and pronociception simultaneously in the rat paw formalin test. Eur J Pharmacol 1993; 236:193-199. [CrossRef]

(19.) Sierralta F Pinardi G, Mendez M, Miranda HF Interreaction of opiods with antidepressantinduced antinociception. Psychopharmacology 1995; 122:374-378. [CrossRef]

(20.) Zurek JR, Nadeson R, Goodchild C. Spinal and supraspinal components of opioid antinociception in streptozotoc in induced diabetic neuropathy in rats. Pain 2001; 90:57-63. [CrossRef]

(21.) Godfroy F Butler SH, Weil-Fugazza J, Besson JM. Do acute or chronic tricyclic antidepresants modify morphine antinociception in rats? Pain 1986; 25:233-244. [CrossRef]

(22.) Yaksh TL. Spinal opiate analgesia: Characteristics and principles of action. Pain 1981; 11:293-346. [CrossRef]

(23.) Schreiber S, Backer MM, Pick CG. The antinociceptive effect of venlafaxine in mice is mediated through both opioid and serotonergic mechanisms. Neurosci Lett 1999; 273:85-88. [CrossRef]

(24.) Gray AM, Pache DM, Sewell RD. Do alpha 2-adrenoceptors play an integral role in the antinociceptive mechanism of action of antidepressant compounds? Eur J Phar 1999; 378:161-168. [CrossRef]

(25.) Ghelardini C, Baleotti N, Bartolini A. Antinociception induced by amitriptyline and imipramine is mediated by alpha2A-adrenoceptors. Jpn J Pharmacol 2000; 82:130-137. [CrossRef]

Ahmet INAL [1], Murat BUYUKSEKERCI [2], Hasan Basri ULUSOY [1]

[1] Department of Pharmacology, Erciyes University School of Medicine, Kayseri, Turkey

[2] Department of Drug and Pharmaceuticals, Ankara Health Directorate, Ankara, Turkey

Correspondence Address: Ahmet inal, Erciyes Universitesi Tip Fakultesi, Farmakoloji Anabilim Dali, Kayseri, Turkiye E-mail:

Received: 14.05.2014 Accepted: 09.03.2015

Table 1. Drugs and doses of the drugs administered to
rats with the development of hyperalgesia

Experimental        Drugs and doses           Rats with
                                           of hyperalgesia
Group 1              Control (ps)                 4
Group 2           Mirtazapine 5 mg/kg             5
Group 3          Mirtazapine 10 mg/kg             4
Group 4          Mirtazapine 15 mg/kg             6
Group 5            Naloxone 1 mg/kg               4
Group 6           Metergoline 2 mg/kg             5
Group 7            BRL 44408 4 mg/kg              4
Group 8             Mirtazapine 15                6
                mg/kg+Nalokson 1 mg/kg
Group 9             Mirtazapine 15                5
               mg/kg+Metergoline 2 mg/kg
Group 10            Mirtazapine 15                4
                mg/kg+BRL 44408 4 mg/kg
Group 11        Streptozotocin 50 mg/kg           4

ps: physiological saline

Table 2. Venous blood glucose levels prior to and after
(3 days) STZ administration in rats; moreover, the body
weights of the animals prior to and 21 days after STZ

                       Before STZ            After STZ
                     (Mean and SE)         (Mean and SE)

Venous blood       1 15 [+ or -] 1.12   384 [+ or -] 1.23 *
  glucose levels
body weights (g)   186 [+ or -] 1.24    145 [+ or -] 1.35 *

* p < 0.05. STZ: streptozotocin; SE: standard error

Table 3. Hot-plate measurement values prior
to and 3 weeks after STZ administration

Group         Hot-plate values
             (s) (Mean and SE)

Before STZ    21 [+ or -] 3.21
After STZ    17 [+ or -] 2.43 *

* p<0.05. SE: standard error; STZ: streptozotocin

Table 4. Effects of drugs on sensorimotor performance

Drug (mg/kg)      Percentage of         Time spent
                 animals that can         on the
                 stand for 2 min       rotarod (s)
                  on the rotarod      (Mean and SE)

Control                100          120 [+ or -] 1.8 *
Mirtazapine 5          100           120 [+ or -] 0 *
Mirtazapine 10          97           118 [+ or -] 1 *
Mirtazapine 15          91           117 [+ or -] 3 *
Naloxone 1             100           120 [+ or -] 0 *
Metergoline 2           98          119 [+ or -] 2.5 *
BRL44408 4              94          112 [+ or -] 3.1 *

* No significant difference was found
between the groups. SE: standard error
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Article Details
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Title Annotation:Research Article
Author:Inal, Ahmet; Buyuksekerci, Murat; Ulusoy, Hasan Basri
Publication:Archives of Neuropsychiatry
Article Type:Report
Geographic Code:7TURK
Date:Mar 1, 2016
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