ARTICLE IN PRESS

The Veterinary Journal ■■ (2014) ■■–■■

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The Veterinary Journal

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Short Communication

Evaluation of three medetomidine-based protocols for chemical

restraint and sedation for non-painful procedures in companion rats
(Rattus norvegicus)
Luca Bellini, Tommaso Banzato, Barbara Contiero, Alessandro Zotti *
Department of Animal Medicine, Production and Health, Clinical Section, University of Padua, Viale dell’Università 16, AGRIPOLIS, Legnaro, Padua 35020,
Italy

A R T I C L E I N F O A B S T R A C T

Article history: Three medetomidine-based protocols were evaluated for sedation in companion rats undergoing diag-
Accepted 24 March 2014 nostic procedures. Group Me received medetomidine at 150 μg/kg intramuscularly (IM); group Me-Bu
received medetomidine 100 μg/kg IM and butorphanol 2 mg/kg IM, and group Me-Bu-Mi received
Keywords: medetomidine 50 μg/kg IM, butorphanol 2 mg/kg IM and midazolam 1 mg/kg IM.
Butorphanol The righting reflex disappeared more quickly in the Me-Bu-Mi group, but recovery after atipamezole
Medetomidine
was longer. In group Me, a palpebral reflex was present throughout sedation in more rats than in the other
Midazolam
two groups. Pulse and respiratory rates were higher when lower doses of medetomidine were used, al-
Rats
Sedation though arterial haemoglobin O2 saturation was similar among groups. All protocols tested produced ad-
equate sedation lasting 25 min.
© 2014 Elsevier Ltd. All rights reserved.

Medetomidine produces poor sedation in rats unless used at high
doses, which can cause cardiovascular and respiratory depression
(Hauptman et al., 2003). In other species, midazolam and
butorphanol are co-administered to decrease the dose of
medetomidine or improve sedation (Sakaguchi et al., 1992; Itamoto
et al., 2000; Kuo and Keegan, 2004). The aim of this study was to
compare the sedation produced by medetomidine (Domitor, Orion
Corporation) administered alone, medetomidine combined with
butorphanol (Dolorex, Intervet Italia), or both drugs combined with
midazolam (Midazolam, Hameln Pharmaceutical).
The Ethics Committee of the University of Padua approved this
study (Protocol 41060; 12 July 2010) and prior owner consent was
obtained. Thirty clinically healthy adult companion rats (Rattus
norvegicus) of both genders were enrolled in a randomised, operator-
blind design. Three groups of 10 rats each were sedated using an
intramuscular (IM) injection in the quadriceps with (1)
medetomidine 150 μg/kg (Group Me); (2) medetomidine 100 μg/
kg and butorphanol 2 mg/kg (Group Me-Bu), or (3) medetomidine
50 μg/kg, butorphanol 2 mg/kg and midazolam 1 mg/kg (Group Me-
Bu-Mi). Sedation was induced so that whole-body ventrodorsal and
lateral radiographs and abdominal ultrasonographic examinations
could be performed. The rats breathed room air throughout the entire
study period.
Sixty seconds after IM injection, the rats were gently rotated on
the flank to evaluate righting reflex (RiRef). The procedure was
* Corresponding author. Tel.: +39 04982 72509.
E-mail address: alessandro.zotti@unipd.it (A. Zotti).
repeated at 30 s intervals until the reflex was lost, or earlier if muscle
relaxation was observed. Radiographs were taken. Five minutes later,
the rats were moved to the ultrasound room and placed in dorsal
recumbency on an electric heating pad (Pet-Mat, Dale Ecotech). Rectal
temperature was measured (Temp0) before the abdomen was clipped.
A pulse oximeter ear probe was placed on the metatarsal region
(Cardiocap, Datex Omeda) and arterial haemoglobin O2 saturation
(SpO2) was recorded. Respiratory rate (RR) was measured by count-
ing chest or abdominal wall movements over 1 min while the pulse
rate (PR) was measured by pulse wave count. Palpebral reflex (PaRef)
was evaluated by touching the nasal canthus of the eye with a cotton
swab. These parameters were recorded every 5 min.
After 20 min, an ultrasound scan was completed, rectal temper-
ature was measured again (Temprec) and atipamezole (Antisedan,
Orion Corporation) was administered subcutaneously (SC)
in the flank region at a dose 2.5 times higher than that of
medetomidine. As rats regained the RiRef, they were moved to a cage
and observed for 1 h to monitor for signs of sedation or abnormal
behaviour.
Data were analysed statistically using a commercially available
statistical package (SAS 9.1, SAS Institute). Normal distribution was
confirmed by a Shapiro–Wilk test, equality of variances by Levene’s
test. Random allocation of animals among groups minimised the risk
of allocation bias. Differences among groups for loss and return of
RiRef, Temp0 and Temprec were analysed with an ANCOVA linear model
with gender, protocol and their interaction as fixed effects and
bodyweight as covariates. A linear repeated mixed model was used
to analyse PR, RR and SpO2 over time. Gender, sedation protocol and

http://dx.doi.org/10.1016/j.tvjl.2014.03.024
1090-0233/© 2014 Elsevier Ltd. All rights reserved.

Please cite this article in press as: Luca Bellini, Tommaso Banzato, Barbara Contiero, Alessandro Zotti, Evaluation of three medetomidine-based protocols for chemical
restraint and sedation for non-painful procedures in companion rats (Rattus norvegicus), The Veterinary Journal (2014), doi: 10.1016/j.tvjl.2014.03.024
 ARTICLE IN PRESS
2 L. Bellini et al./The Veterinary Journal ■■ (2014) ■■–■■

Table 1
Results of the ANCOVA linear model analysis of the effects of three medetomidine-based sedation protocols.

Group (mean ± standard deviation) Sedation protocol Bodyweight Gender Gender x sedation
protocol

Me Me-Bu Me-Bu-Mi F-stat P F-stat P F-stat P F-stat P

Time to loss of RiRef (s) 543 ± 271 426 ± 235 131 ± 42a 9.69 <0.001 0.61 0.442 0.20 0.662 1.19 0.321
Time to return of RiRef (s) 190 ± 62 172 ± 54 332 ± 41a 12.01 <0.001 5.88 0.024 0.04 0.853 1.78 0.191
Temp0 (°C) 34.6 ± 1.0 34.8 ± 0.9 34.8 ± 0.9 0.34 0.718 0.80 0.382 2.79 0.112 0.85 0.444
Temprec (°C) 34.6 ± 1.0 35.0 ± 0.5 35.1 ± 0.1 1.21 0.317 3.13 0.091 2.46 0.131 0.07 0.935

Me, rats that received 150 μg/kg IM medetomidine; Me-Bu, rats that received 100 μg/kg medetomidine IM and 2 mg/kg butorphanol IM; Me-Bu-Mi, rats that received 50 μg/
kg medetomidine IM, 1 mg/kg midazolam IM and 2 mg/kg butorphanol IM; RiRef, righting reflex; Temp0, temperature at the beginning of the ultrasonographic exam; Temprec,
temperature before atipamezole.
a Statistically significant differences between group Me-Bu-Mi and groups Me-Bu and Me (P < 0.001).

their interaction were inserted as fixed effects, bodyweight was a
covariate, and time and animal effects were included as repeated
and random effects respectively. Interaction between sedation pro-
tocol and time was also inserted in the model and the Bonferroni
post-hoc pair wise comparison test was used if statistically signif-
icant differences were detected. The number of animals with a pos-
itive palpebral reflex was compared at each time point using Fisher’s
exact test. P < 0.05 was considered statistically significant.
There were no adverse events recorded in any of the groups. The
RiRef was lost earlier in group Me-Bu-Mi than in group Me or group
Me-Bu, but it reappeared later; this effect was inversely associated
with bodyweight (Table 1). Temp0 and Temprec were similar among
groups (Table 1). The effects of gender, sedation protocol, their in-
teraction and bodyweight on Temp0 and Temprec are shown in Table 1.
The palpebral reflex was maintained in >50% of rats in group Me
(Fig. 1a).

Fig. 1. Trends over time for rats sedated for diagnostic procedures with medetomidine alone at 150 μg/kg IM (Me) or with medetomidine 100 μg/kg and butorphanol 2 mg/
kg IM (Me-Bu), or medetomidine 50 μg/kg, midazolam 1 mg/kg and butorphanol 2 mg/kg IM (Me-Bu-Mi). (a) Number of rats with a positive palpebral reflex. Points with
different letters are statistically different (P < 0.05). (b) Pulse rate, (c) respiratory rate and (d) arterial haemoglobin O2 saturation in the three groups of rats. Data represent
mean ± standard deviation. (#) Statistically significant difference between Me-Bu-Mi and both Me-Bu and Me (P < 0.05). (*) Statistically significant difference between Me-
Bu-Mi and Me (P < 0.05).

Please cite this article in press as: Luca Bellini, Tommaso Banzato, Barbara Contiero, Alessandro Zotti, Evaluation of three medetomidine-based protocols for chemical
restraint and sedation for non-painful procedures in companion rats (Rattus norvegicus), The Veterinary Journal (2014), doi: 10.1016/j.tvjl.2014.03.024
 ARTICLE IN PRESS
L. Bellini et al./The Veterinary Journal ■■ (2014) ■■–■■ 3

Table 2
Results of the linear repeated mixed model for the effects of the sedation protocol, time, their interaction, gender, gender and sedation interaction, bodyweight and the animal
effect expressed as percentage of total variation in heart rate, respiratory rate and arterial O2 saturation.

Pulse rate Respiratory rate Arterial haemoglobin

O2 saturation

F-stat P F-stat P F-stat P

Sedation protocol 3.97 <0.001 3.84 0.037 0.01 0.994

Time 5.29 0.033 0.17 0.952 1.86 0.124
Sedation protocol x time 0.84 0.568 2.57 <0.014 0.67 0.718
Gender <0.01 0.985 0.12 0.735 0.60 0.445
Gender × sedation protocol 0.71 0.500 0.15 0.861 0.83 0.447
Bodyweight <0.01 0.984 0.07 0.794 0.20 0.657
Animal effect (% of total variation) 69 87 39

Rats in group Me had lower mean PR compared to those in
group Me-Bu; PR decreased over time regardless of the sedation
protocol used (Fig. 1b; Table 2). There was a statistically signifi-
cant effect of the protocol on the RR; this parameter was higher in
group Me-Bu-Mi and group Me-Bu than in group Me (Fig. 1c;
Table 2). In group Me, RR was statistically lower than in group Me-
Bu-Mi at 0, 15, and 20 min (Fig. 1c). No differences in SpO2 were de-
tected among sedation protocols or over time (Fig. 1d; Table 2). The
linear repeated mixed model did not demonstrate the influence of
other effects on PR, RR or arterial O2 saturation (Table 2).
Based on the results of this study, all of the protocols tested pro-
vided sufficient sedation to perform non-painful imaging proce-
dures in companion rats. The onset of sedation was faster if
midazolam was added to medetomidine and butorphanol, despite
a lower dose of medetomidine. In this study, as observed in other
species, butorphanol added to medetomidine prolonged the dura-
tion of medetomidine sedation, with minimal effects on the time
required to reach lateral recumbency (Sakaguchi et al., 1992; Kuo
and Keegan, 2004). Midazolam acts synergistically with α2 recep-
tor agonists such as medetomidine, shifting the dose–response curve
to the left; this might explain the relatively short time before the
RiRef was lost and the complete suppression of the PaRef in this
group (Salonen et al., 1992). The sedative effect of midazolam could
have contributed to the delay in the return of the RiRef after
atipamezole.
Medetomidine produced a dose-dependent decrease in PR and
RR, as previously described in rats and other species, regardless of
the co-administration of midazolam and butorphanol (Sakaguchi
et al., 1992; Hauptman et al., 2003). The rats breathed room air during
sedation in this study, but O2 supplementation should be consid-
ered, as SpO2 values <90% were recorded regardless of the proto-
col used. The rats also became hypothermic during sedation despite
the use of a heating pad; therefore care should be taken during re-
covery to avoid any further decrease in body temperature.
In conclusion, all three protocols can be used for sedation in com-
panion rats, but the fastest onset of sedation was achieved when
midazolam was combined with medetomidine and butorphanol. The
use of medetomidine alone produced lower mean PR and RR than
the other protocols.
Conflict of interest statement
None of the authors of this study has any financial or personal
relationship with people or organisations that could inappropri-
ately influence or bias the contents of this paper.
References
Hauptman, K., Jekl, V., Knotek, Z., 2003. Use of medetomidine for sedation in the
laboratory rat (Rattus norvegicus). Acta Veterinaria Brno 72, 583–591.
Itamoto, K., Hikasa, Y., Sakonjyu, I., Itoh, H., Kakuta, T., Takase, K., 2000. Anaesthetic
and cardiopulmonary effects of balanced anaesthesia with medetomidine-
midazolam and butorphanol in dogs. Journal of Veterinary Medicine. A,
Physiology, Pathology, Clinical Medicine 47, 411–420.
Kuo, W.C., Keegan, R.D., 2004. Comparative cardiovascular, analgesic, and sedative
effects of medetomidine, medetomidine-hydromorphone, and medetomidine-
butorphanol in dogs. American Journal of Veterinary Research 65, 931–937.
Sakaguchi, M., Nishimura, R., Sasaki, N., Ishiguro, T., Tamura, H., Takeuchi, A., 1992.
Enhancing effect of butorphanol on medetomidine-induced sedation in pigs. The
Journal of Veterinary Medical Science 54, 1183–1185.
Salonen, M., Reid, K., Maze, M., 1992. Synergistic interaction between alpha
2-adrenergic agonists and benzodiazepines in rats. Anesthesiology 76, 1004–
1011.

Please cite this article in press as: Luca Bellini, Tommaso Banzato, Barbara Contiero, Alessandro Zotti, Evaluation of three medetomidine-based protocols for chemical
restraint and sedation for non-painful procedures in companion rats (Rattus norvegicus), The Veterinary Journal (2014), doi: 10.1016/j.tvjl.2014.03.024