L5 Lumbar Laminectomy in Aged C57BL/6J Mice to Model Postoperative Cognitive Dysfunction

Summary

This protocol describes a standardized L5 lumbar laminectomy in aged C57BL/6J mice to establish a model of postoperative cognitive dysfunction while minimizing locomotor impairment, thereby supporting behavioral, histological, and mechanistic studies of perioperative neurocognitive disorders.

Abstract

Postoperative cognitive dysfunction (POCD) is a common neurological complication following surgery and anesthesia, particularly in older adults, and is associated with delayed recovery, prolonged hospitalization, and reduced quality of life. Orthopedic and spinal procedures are linked to a relatively high risk of POCD; however, preclinical models that closely reflect the perioperative features of spinal surgery remain limited. Commonly used models, including laparotomy and tibial fracture, may produce persistent locomotor impairment and prolonged peripheral inflammation that can confound the interpretation of cognitive behavioral outcomes. Here, this protocol describes a standardized, low-cost protocol for modeling POCD using L5 lumbar laminectomy under sevoflurane anesthesia in 18-month-old male C57BL/6J mice. The procedure is performed using routine surgical instruments and produces a controlled surgical injury while minimizing persistent motor deficits. Cognitive function is evaluated using the novel object recognition test, novel arm Y-maze, and Barnes maze, whereas hippocampal neuronal injury and neuroinflammatory changes are assessed by Nissl staining and Iba1 immunofluorescence, respectively. Representative results demonstrate preserved locomotor activity, impaired cognitive performance, reduced hippocampal neuronal integrity, and enhanced hippocampal neuroinflammation following surgery. This standardized and accessible model provides a clinically relevant platform for investigating the mechanisms underlying spine surgery-associated cognitive dysfunction and for evaluating candidate perioperative neuroprotective interventions.

Introduction

Postoperative cognitive dysfunction (POCD) is defined as a decline in learning and memory, attention, processing speed, and executive function after anesthesia and surgery¹, predominantly affecting elderly patients, and its reported incidence appears to vary by surgical context and patient population^2,3. POCD has been reported to occur frequently among older adults undergoing elective orthopedic surgery⁴. Clinically, POCD may persist for weeks to months after surgery and has been associated with prolonged hospitalization and increased mortality^5....

Protocol

All animal experiments were performed in accordance with the Guide for the Care and Use of Laboratory Animals and approved by the Animal Care and Use Committee of Zhejiang University (Ethics approval number: ZJU20260153).

1. Preoperative preparation

1. Experimental animals, group allocation, analgesia, and blinding
   1. Use 18-month-old male C57BL/6J mice weighing 30–34 g. House the mice under a 12 h light/dark cycle with free access to food and water.
   2. Allow the mice to acclimate to the housing conditions for at least 7 days before surgery. Randomly allocate the mice to the L5 laminectomy group or....

Results

All mice survived to the scheduled experimental endpoint, none met the predefined humane endpoint criteria, and all were included in the final analyses. Motor function was evaluated on postoperative day 3 using open-field and gait analyses. Neither total distance traveled nor average speed in the open-field test differed between groups, indicating comparable spontaneous locomotor activity (Figure 2A). Gait analysis likewise showed no significant between-group differences in stride length, st.......

Discussion

Postoperative cognitive dysfunction has been reported in older adults undergoing spinal surgery¹⁰. Experimental evidence further indicates that aging increases susceptibility to surgery-associated cognitive impairment and neuroinflammatory responses^11,12. The present protocol combines approximately 4 h of sevoflurane anesthesia with L5 laminectomy in aged mice to model early postoperative cognitive impairment. Its principal methodological .......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
C57BL/6J male mice	Suzhou Industrial Park Ai'er Maite Technology Co., Ltd.	SYXK2023-0005	Animal model
ANY-maze Video Tracking Software	Stoelting Co., USA	6000	Behavioral video tracking and analysis
Erythromycin ophthalmic ointment	Beijing Twinluck Pharmaceutical Co,. Ltd.	H11021270	Prevent eye dry
Fine forceps	NA	NA	Clip the skin
Heating pad	NA	NA	Temperature-controlled
Iba1/AIF-1 (E4O4W) Rabbit Monoclonal Antibody	Cell Signaling Technology	17198	Iba1 immunofluorescence staining
Inhalant anesthesia equipment	RWD Life Science Co., Ltd.	R581S	Anesthesia
MGT-PR	Anhui Zhenghua Biological Instrument Equipment Co., Ltd., China	MJY2025112704	Gait analysis
Micro Ophthalmic scissors	NA	NA	Bone removal
Nissl staining solution	Haoke Biotechnology Co., Ltd.	HK2033	Nissl staining
Scissors	Shang Shou Jin Zhong Surgical Instruments Jiangsu ITD.,Corp	Y00030	Skin incision
Sevoflurane	Shanghai Hengrui Pharmaceutical Co,. Ltd.	25062331	Anesthesia
Single Use Surgical Drape	Jiangxi Zhizheng Medical Equipment Co., Ltd.	20212140221	Platform for surgical operation
Small animal trimmer	Yiwu Leiben Trading Co., Ltd.	2504KFZB3592-03310	Hair removal
Surgical Suture Needles with Thread	Shanghai Pudong Jinhuan Medical Products Co., LTD.	F603	Suture skin
Veet hair removal cream	RECKITT BENCKISER (India) Ltd	NA	Hair removal