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Article 

Anaesthetic Efficacies of Epidurally Administered Lignocaine-
HCl and Bupivacaine-HCl Alone and Their Combination in 
Rabbits 
Roshik Shrestha 1*, Rajesh Tharu 2, Jyoti Chaudhary 1, Flavia Pradhan 1, Dibina Kaini 1, Shashank Shrestha 1, and 
Egendra Shrestha 3 

1 Nepal Polytechnic Institute; sthroshik@gmail.com (R.S.), jyotichau147@gmail.com (J.C), flaviapra-
dhan7@gmail.com (F.P), (S.S) 

2 Nepal Polytechnic Institute; Faculty of Veterinary Surgery and Radiology; raaztharu83@gmail.com (R.T.) 
3   Nepal Polytechnic Institute; Research Director; egendrastha@gmail.com (E.S.) 
* Correspondence: sthroshik@gmail.com 

Abstract: The present study aimed to evaluate the anaesthetic efficacy of epidurally administered lignocaine-HCl and bupivacaine-
HCl, both individually and in combinations, in rabbits. A total of 15 rabbits was equally divided into three groups. Group A received 
an epidural injection of 4 mg/kg of 2% lignocaine-HCl, while Group B received 1 mg/kg of 0.5% bupivacaine-HCl. Group C was 
administered a combined solution of 2 mg/kg of 2% lignocaine-HCl and 0.5 mg/kg of 0.5% bupivacaine-HCl. Physiological 
parameters such as heart rate, respiratory rate, and rectal temperature were recorded 10 minutes before and then at 10-minute 
intervals for 120 minutes post-epidural anaesthesia. Additionally, the onset and duration of anaesthesia, onset and duration of loss 
of weight-bearing ability, and onset and duration of flaccid paralysis were observed following epidural administration. The 
anaesthesia onset times were 8.0 ± 0.354 minutes, 12.5 ± 0.224 minutes, and 10.1 ± 0.272 minutes in rabbits receiving lignocaine, 
bupivacaine, and the lignocaine + bupivacaine combination, respectively. The duration of anaesthesia was significantly higher (P < 
0.01) in Group B (138.00 ± 5.15 minutes) than in Group A (50.60 ± 1.60 minutes) and Group C (87.20 ± 5.05 minutes). The onset and 
duration of loss of weight-bearing ability and flaccid paralysis were significantly higher (P < 0.05) in Group B (17.60 ± 0.245 minutes) 
compared to Groups A and C. The lumbosacral epidural administration of the combined lignocaine and bupivacaine solution 
provided enhanced anaesthetic effects compared to lignocaine or bupivacaine alone. 

Keywords: Analgesia; Regional anesthetics; Lumbosacral; Rabbits 

 
 

1. Introduction 

Lumbosacral regional anesthesia results in effective analgesia due to its proximity to the 
spinal cord receptors responsible for the regulation and transmission of nociceptive 
signals [1], [2]. The epidural administration of regional anesthetic is relatively safe and 
provides effective anesthesia and post-operative analgesia. The drugs and their 
combinations have been used to induce epidural analgesia in dogs and cats [3], [4],[5]. 
The ideal epidural local anesthetic should have a rapid onset of action, a long duration of 
action, good analgesia, and muscular relaxation [6]. There is no single anesthetic that 
possesses all of these qualities. Although lignocaine has a short latency period for epidural 
anesthesia, its effectiveness becomes limited for longer surgical procedures [7]. 
Bupivacaine lasts longer but it takes more time to onset and its muscular relaxation is also 
poor [6]–[8]. Therefore, an agent with a rapid onset and a sufficient duration of action 
would be ideal and could be obtained with regional anesthetic mixtures that combine the 

Received: 09.10.2024 

Accepted: 20.11.2024 

Published: 31.12.2024 

DOI: 10.52331/v29i4g55 

 

 

 

Copyright: © 2024 by the authors. 

Submitted for possible open access 

publication under the terms and 

conditions of the Creative Commons 

Attribution (CC BY) license 

(http://creativecommons.org/licenses

/by/4.0/). 

mailto:sthroshik@gmail.com
mailto:jyotichau147@gmail.com
mailto:flaviapradhan7@gmail.com
mailto:flaviapradhan7@gmail.com
mailto:raaztharu83@gmail.com
mailto:egendrastha@gmail.com


Cluj Vet J 2024, vol 29, issue 4 20 of 38 
 

 

desirable properties of each component drug. To the knowledge of the authors, there are no studies of 
epidural regional anesthesia using a combination of lignocaine and bupivacaine in rabbits. 

The present study aimed to compare the effects of lignocaine and bupivacaine alone and their combinations 
for epidural anesthesia considering the onset and duration of analgesia in rabbits.   

 
2. Materials and Methods 

Study Location 

This study was done from August- 2023 to October- 2023 in the NPI college research unit  

Experimental Rabbits 

Fifteen clinically normal rabbits (Oryctolagus cuniculus) weighing 1.8 ± 2.3 kg were used in this study. They 
were purchased from a local rabbit market in Bharatpur.  The rabbits were housed in a 12-h light-dark 
photoperiod in 3 different cages of the research laboratory of NPI College. They were fed seasonal grasses, 
concentrates, and water ad libitum. The rabbits were kept for 2 weeks for acclimatization in in laboratory 
environment. The experimental protocol was approved by the Nepal Veterinary Council on animal research 
(Ref. no. 35/2080/81). 

Study design 

Food was withheld from the rabbits for 8 hours (i.e. overnight) prior to the experiments, while water was 
allowed freely. The body weight of each rabbit was determined using a weighing balance. The rabbits were 
restrained to facilitate the epidural anesthesia. The injection site was located and the hair of the lumbosacral 
region was shaved and the skin was disinfected. Then after, 2% lignocaine gel was applied topically on site 
to reduce pain and to prevent unnecessary suffering and the epidural puncture was performed using 26 
gauge*5/8” needle. A total of fifteen rabbits were divided into three groups, with each group containing five 
different rabbits: the LIG group, the BUP group, and the LIG+BUP group. The rabbits of LIG, BUP, and 
LIG+BUP groups received the lumbosacral epidural administration of 2% lignocaine (4 mg/kg) 0.5% 
bupivacaine (1mg/kg) and 1:1 mixture of 2% lignocaine and 0.5% bupivacaine (2 mg/kg and 0.5 mg/kg). The 
dose rate were used in this study following  the doses of  prior studies [9]. Physiological parameters were 
recorded 10 minutes before the epidural anesthesia and then after anesthesia at every 10 min intervals over 
a period of 120 min. 

Assessment of Epidural Anesthesia 

The onset of anesthesia was assessed by observing the reflexes to needle pinprick stimulation to the hind 
paddle or the web. It was performed at one-minute intervals until surgical anaesthesia. However, the 
duration of anesthesia was determined by the return of response to a bipedal pinprick stimulus. Loss of 
weight-bearing was detected when rabbits did not stand on its hind limbs. The recovery was confirmed 
when the rabbits became stable. Flaccid paralysis was confirmed when no measurable tone in both hind 
limbs. The heart rate (HR), respiratory rate (RR) and rectal temperature (RT) were recorded appropriately 
10 minutes before and then after every 10 minutes interval over a period of 120 min of epidural injections of 
regional anesthetics.   

Statistical analysis 



Cluj Vet J 2024, vol 29, issue 4 21 of 38 
 

 

The data are expressed as Mean ± Standard Error of Mean. The data was analyzed in Minitab version 17 

software manufactured in Pennsylvania State University. Data were analyzed using one-way ANOVA fol-

lowed by post hoc Tukey’s test. However, the physiological variables were compared using for repeated 

measures in each group. The P value less than 0.05 was considered significant. 

3. Results 

The onset of anesthesia with LIG-Group (8.0 ± 0.354 min), BUP-Group (12.5 ± 0.224 min) and LIG-BUP 

combination- Group (10.1 ± 0.272 min) was significantly different (P<0.05) from each other among the three 

groups. The mean duration of anesthesia of LIG group (50.60 ± 1.60), BUP group (138.00 ± 5.15), and 

LIG+BUP group (87.20 ± 5.05) differs highly significantly (P<0.01) from each other. The mean onset of loss 

of weight-bearing ability was significant (P<0.05) in the BUP group with (17.6000 ± 0.245). The mean duration 

of loss of weight-bearing ability was highly significant (P<0.05) in the BUP group (76.80 ± 3.57) compared to 

LIG group and LIG+BUP which also differs from each other significantly (P<0.05) with values of (24.000 ± 

0.949) and (49.00 ± 2.74). The mean onset of flaccid paresis in BUP group (19.600 ± 0.927) was highly 

significant (P<0.05) as compared to LIG group and LIG+BUP group with values of (23.900 ± 0.400) and 

(24.600 ± 0.927) respectively. The mean duration of flaccid paresis was highly significant (P<0.05) in LIG 

group (10.900 ± 0.332) compared to the BUP group and LIG+BUP group which also differs from each other 

significantly (P<0.05) with values of (50.20 ± 2.22) and (19.00 ± 1.52) respectively. There were no significant 

changes in heart rate, respiration rate and temperature. 

Table 1: The mean ± SEM of onset time of anesthesia (min), duration of anesthesia (min), onset and 
duration of loss of weight bearing ability (min), onset and duration of flaccid paresis (min). 

Parameter LIG BUP LIG+BUP 

Time of onset of anesthesia  8 ± 0.354 a 12.5 ± 0.224 b 10.1 ± 0.272 c 

Duration of anesthetic effect  50.60 ± 1.60 a 138.00 ± 5.15 b 87.20 ± 5.05 c 

Onset of loss of weight-
bearing ability 

16.00 ± 0.707 a 17.60 ± 0.245 b 16.00 ± 0.274 a 

Duration of loss of weight 
bearing ability 

24.00 ± 0.949 a 76.80 ± 3.57 b 49.00 ± 2.74 c 

Onset of  flaccid paralysis 23.90 ± 0.400 a 19.60 ± 0.927 b 24.60 ± 0.927 a 

Duration of flaccid paresis 10.90 ± 0.332 a 50.20 ± 2.22 b 19.00 ± 1.52 c 

    Figures with similar symbols in the same row indicate non- significant 

 

 

Table 2: The mean ± SEM of HR (beat/min) measured at different times 10 minutes prior and then after 
anesthesia at every 10 minute intervals over a period of 120 min post injection. 

Time LIG BUP LIG+BUP 

-10 232.60 ± 9.79 225.4 ± 10.3 222.80 ± 4.79 

10 218.60 ± 7.61 242.8 ± 15.2 215.60 ± 6.18 



Cluj Vet J 2024, vol 29, issue 4 22 of 38 
 

 

20 200.40 ± 8.18 213.80 ± 9.68 200.00 ± 5.67 

30 208.40 ± 7.28 206.2 ± 10.8 208.40 ± 5.61 

40 234.40 ± 6.76 203.2 ± 13.8 210.00 ± 7.13 

50 225.2 ± 11.9 197.4 ± 12.1 208.40 ± 3.82 

60 203.6 ± 12.3 204.60 ± 9.91 204.40 ± 2.99 

70 203.4 ± 13.2 205.00 ± 7.63 221.60 ± 2.64 

80 218.80 ± 7.71 209.80 ± 8.32 212.2 ± 10.1 

90 216.40 ± 4.30 210.00 ± 6.88 218.40 ± 2.66 

100 215.80 ± 4.61 203.60 ± 8.19 221.60 ± 3.06 

110 219.80 ± 6.00 207.6 ± 13.7 223.20 ± 1.53 

120 214.00 ± 6.03 211.4 ± 11.1 225.20 ± 2.71 

 

Table 3. The mean ± SEM of RR (breaths/min) measured at different times 10 minutes prior and then after 
anesthesia at every 10 minute intervals over a period of 120 min post injection 

Time LIG BUP LIG+BUP 

-10 129.60 ± 3.83 148.6 ± 11.8 143.2 ± 12.6 

10 127.40 ± 4.49 159.6 ± 7.46 135.20 ± 6.21  

20 123.40 ± 7.70 147.40 ± 7.40 141.40 ± 6.66 

30 125.00 ± 7.30 138.4 ± 10.2 144.20 ± 6.33 

40 122.6 ± 13.2 142.6 ± 10.9 145.20 ± 9.09 

50 131.2 ± 14.6 134.0 ± 12.0 142.80 ± 8.55 

60 142.4 ± 16.0 137.8 ± 12.6 149.60 ± 9.10 

70 149.8 ± 15.5 136.8 ± 10.8 138.40 ± 7.09 

80 140.0 ± 12.7 143.0 ± 12.3 146.60 ± 7.24 

90 138.4 ± 11.0 146.6 ± 15.3 135.60 ± 7.30 

100 134.20 ± 8.01 144.0 ± 17.2 135.60 ± 6.56 

110 131.60 ± 9.05 146.4 ± 13.3 139.60 ± 8.42 

120 129.40 ± 6.46 149.0 ± 10.8 137.00 ± 6.71 

 

Table 4. . The mean ± SEM of RT (°F) measured at different times 10 minutes prior and then after 
anesthesia at every 10 minutes intervals over a period of 120 min post injection 

Time LIG BUP LIG+BUP 

-10 101.76 ± 0.129 101.68 ± 0.0583 101.84 ± 0.211 

10 101.56 ± 0.0510  101.46 ± 0.0872  101.60 ± 0.0600  

20 101.54 ± 0.0510 101.32 ± 0.0735 101.42 ± 0.121 

30 101.56 ± 0.103 101.40 ± 0.268 101.52 ± 0.0583 

40 101.60 ± 0.0707 101.30 ± 0.103 101.36 ± 0.354 

50 101.72 ± 0.116 101.34 ± 0.103 101.42 ± 0.0840 



Cluj Vet J 2024, vol 29, issue 4 23 of 38 
 

 

60 101.64 ± 0.0748 101.38 ± 0.0800 101.32 ± 0.371 

70 101.50 ± 0.0707 101.42 ± 0.180 101.46 ± 0.0812 

80 101.40 ± 0.0949 101.34 ± 0.121 101.34 ± 0.206 

90 101.70 ± 0.210 101.38 ± 0.185 101.38 ± 0.273 

100 101.7 ± 0.169 101.40 ± 0.0949 101.70 ± 0.0949 

110 101.65 ± 0.150 101.48 ± 0.107 101.52 ± 0.0663 

120 101.68 ± 0.133 101.48 ± 0.0374 101.50 ± 0.134 

 

4. Discussion 

None of the rabbits died or showed any side effects during and after anesthesia for a week period. In this 
study, the rabbits were gently restrained for injection of epidural anesthesia with short needles which 
produced no apparent discomforts. The lumbosacral epidural anesthesia technique in rabbits and ferrets 
(Mustela furo) is identical to dogs and cats, with the exception of the rarely definitive popping sensation 
when the intervertebral ligament is punctured [10]. The spinal cord continues caudally into the sacral 
vertebrae in rabbits and thereby increased the risk of puncture of both the dura and arachnoid membranes 
during lumbosacral epidural injection [11]. To overcome this situation, the anesthesia was administered once 
cerebrospinal fluid was seen in the hub of the needle. In this study, solutions of lignocaine and bupivacaine 
were observed to be dispersed in the syringe, showing pharmacological compatibility which is similar to 
previous studies [12]. The doses of LIG 0.2 ml/kg i.e. (4 mg/kg) and BUP 0.2 ml/kg i.e. (1 mg/kg) were used 
in this study following  the doses of  prior studies [9]. The combination of lignocaine and bupivacaine 
exhibited a longer onset of action (P<0.05) than lignocaine but shorter (P<0.05) than that of bupivacaine. This 
result is similar findings by [13]. Onset of action of regional anesthetics differs due to pka (acid dissociation 
constant or pH at which the non-ionized and ionized fractions are at equilibrium) values when pH value of 
tissue remains constant. Regional anesthetics with a low pKa of 7.6-7.9 have a rapid onset of action because 
30% to 40% of these drugs exist in the unionized state at pH 7.4 of which lignocaine has pKa value of 7.9. 
Lignocaine has a faster onset time than drugs with a high pka because its pka is closer to tissue pH [7]. 
Conversely, regional anesthetics with high pKa of 8.0 - 8.9 are slow acting agents because only 15% or less 
of these drugs are unionized at pH 7.4 of which bupivacaine holds value of 8.1.  

Moreover, the duration of analgesia was significantly longer with lignocaine and bupivacaine combination 
(P<0.01) than lignocaine alone and significantly shorter than bupivacaine alone (P<0.01), indicating that 
adding bupivacaine to lignocaine accelerates and prolongs the duration of anesthesia. Similar findings were 
reported in a previous study conducted on dogs [13]. The mean duration of loss of weight bearing ability 
was significantly high in lignocaine- bupivacaine combination group (P<0.05) than in lignocaine alone 
indicating that adding bupivacaine to lignocaine accelerates and prolongs the quality of analgesia. Onset of 
flaccid paresis was significantly rapid (P<0.05) in bupivacaine group. The mean duration of flaccid paresis 
was significantly longer in the lignocaine bupivacaine combination group (P<0.01) than in lignocaine alone. 
This finding can be attributed to a synergistic effect between lignocaine and bupivacaine. The combination 
of the two drugs can decrease the side effects of each drug and increase the duration of flaccid paresis and 
analgesia [14]. No significant differences were observed in heart rate, respiration rate and temperature 
between three groups. 

 



Cluj Vet J 2024, vol 29, issue 4 24 of 38 
 

 

5. Conclusions 

The combination of lignocaine and bupivacaine showed an ideal anesthetic effect over lignocaine or 
bupivacaine alone having shorter onset and prolonged duration of action with good analgesia. None of the 
treatments induced significant changes in heart rate, respiration rate and temperature. Further research is 
needed to investigate the analgesic effect of lignocaine and bupivacaine combination in rabbit under surgical 
conditions. 

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