Chronic Constriction Injury of the Distal Infraorbital Nerve (DIoN-CCI) in Mice to Study Trigeminal Neuropathic Pain

Published: March 08, 2024

doi:

¹Antwerp Surgical Training, Anatomy and Research Centre, Algology (pain research),University of Antwerp, ²Multidisciplinary Pain Center,Antwerp University Hospital

Summary

Chronic constriction injury of the distal infraorbital nerve in mice induces changes in spontaneous behavior (increased face grooming activity) and nocifensive behavior in response to tactile stimulation (hyperresponsiveness to von Frey hair stimulation) that are signs of ongoing pain and allodynia and serves as a model for trigeminal neuropathic pain.

Abstract

Animal models remain necessary tools to study neuropathic pain. This manuscript describes the distal infraorbital nerve chronic constriction injury (DIoN-CCI) model to study trigeminal neuropathic pain in mice. This includes the surgical procedures to perform the chronic constriction injury and the postoperative behavioral tests to evaluate the changes in spontaneous and evoked behavior that are signs of ongoing pain and mechanical allodynia. The methods and behavioral readouts are similar to the infraorbital nerve chronic constriction injury (IoN-CCI) model in rats. However, important changes are necessary for the adaptation of the IoN-CCI model to mice. First, the intra-orbital approach is replaced by a more rostral approach with an incision between the eye and the whisker pad. The IoN is thus ligated distally outside the orbital cavity. Secondly, due to the higher locomotor activity in mice, allowing rats to move freely in small cages is replaced by placing mice in custom-designed and constructed restraining devices. After DIoN ligation, mice exhibit changes in spontaneous behavior and in response to von Frey hair stimulation that are similar to those in IoN-CCI rats, i.e., increased directed face grooming and hyperresponsiveness to von Frey hair stimulation of the IoN territory.

Introduction

Neuropathic pain arises from damage to the somatosensory nervous system, leading to abnormal transmission of sensory signals to the brain. Somatosensory nerve damage does not always lead to neuropathic pain, but the prevalence increases with the severity of clinical neuropathy¹^,². Neuropathic pain patients experience specific symptoms such as spontaneous sensations (burning, pins and needles, electric sensations) and abnormally intense or prolonged pain to innocuous or noxious stimulation that tend to become chronic and resistant to treatment with conventional pain medication³. Significant progress in the field of neuropathic pain research stems from the discovery that loosely constricting ligatures around the sciatic nerve in rats leads to behaviors resembling human neuropathic pain conditions⁴. The animals display reduced thresholds to heat, cold, and mechanical stimulation, and exhibit nocifensive behaviors. Despite the inherent biological differences in pain processing between humans and rodents, animal models are a valuable tool for studying the underlying mechanisms in the development of neuropathic pain and testing newly proposed treatment strategies.

Sensory reflex-based pain testing paradigms have been extensively used in neuropathic pain models, but measuring ongoing pain or other frequently accompanied disturbances (sleeping disorder, depression, anxiety) has not received sufficient attention considering that these are common clinical symptoms affecting quality of life⁵^,⁶^,⁷^,⁸. Face grooming behavior in rats has been documented as a measure of spontaneous neuropathic pain following chronic constriction injury (CCI) of the infraorbital nerve (IoN)⁹^,¹⁰. In addition, rats also develop hyperresponsiveness to mild tactile stimulation of the IoN territory, which is indicative of mechanical allodynia.

Compared to mice, because of their larger size, rats are better suited for surgical injuries. However, mice offer cost and space efficiency and require smaller drug quantities. Also, the advent of transgenic technology has further boosted the use of mice¹¹^,¹². Therefore, the overall goal of this procedure is to perform a surgical infraorbital nerve injury in mice, similar to that in rats, that induces changes in spontaneous and evoked behavior for the study of trigeminal neuropathic pain.

Protocol

Animals are treated and cared for according to the guidelines for pain research in conscious animals of the International Association for the Study of PAIN and in line with the Flemish and European regulations for animal research and the ARRIVE guidelines. The protocol is approved by the institutional Ethical Committee. 1. Animals Use male and female C57BL/6J mice (Janvier, 10 weeks old at arrival). House male and female mice separately in standard solid-…

Representative Results

DIoN-CCI mice show a strong postoperative increase in time spent on isolated face grooming and the number of isolated face grooming episodes (Figure 3). The strongest increase occurs during the first postoperative week and then becomes smaller during the following weeks but is significantly increased for at least 6 weeks. Face grooming during body grooming is more or less unaffected. DIoN-CCI mice are almost completely unresponsive to ipsilateral mechanical stimul…

Discussion

In rats, it has been previously argued that an intra-orbital approach to the IoN is preferable, considering the importance of intact fine musculature controlling complex whisking patterns in vibrissotactile discrimination and the relative distance of the mid-line incision to the cutaneous infraorbital nerve territory¹⁰. Others have argued that a distal approach via an incision into the hairy skin caudal to the vibrissal pad has a number of benefits¹³^,<sup class="…

Divulgaciones

The authors have nothing to disclose.

Acknowledgements

The authors have no acknowledgments.

Materials

Chromic catgut (6-0)	Dynek	CG602D	ligatures
Cotton applicator	Pharmacy
Digital video camera	Sony	HDR-CX330E
Dumont #5 forceps	Fine Science Tools	11251-10
Dumont forceps – Micro-blunted tips (#5/45)	Fine Science Tools	11253-25
Duratears	Alcon	0037-820	ophthalmic ointment
Hooked ligation aid	Fine Science Tools	18062-12
Ketalar	Pfizer		ketamine (50 mg/mL)
Operation microscope	Kaps	SOM 62
Precision cotton swab	Qosina	10225
Precision trimmer	Philips	HP6392/00
Rompun	Bayer		xylazine (2%)
Scissors – blunt tips	Fine Science Tools	14574-09
Semmes-Weinstein Von Frey Aesthesiometer kit	Stoelting	58011
Vicryl Rapide	Ethicon	MPVR489H	sutures

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