The degree of injury decreased with increasing distance from site of injection, with the zone of severe injury (if present) surrounded by zones of intermediate and no injury, in succession (Fig. 3). As the concentration of the LA decreased as it dispersed, a progressive decline in damage away from the injection site was seen. Administration of saline resulted in intermediate damage to nerves, indicating a baseline level of injury associated with injecting any agent into a nerve.
In the negative control group, administration of 50 µL saline resulted in no severe injury to nerve fibers. This is in contrast to bupivacaine, lidocaine, and ropivacaine, where nearly all of the injected nerves showed areas of severe injury, defined by total loss of large myelinated fibers and extensive fibrosis. Regardless of the drug used, there was a significant decrease in number of fibers (bupivacaine P < 0.0001, lidocaine P < 0.0001, and ropivacaine P < 0.0001) and percent nerve in severely injured nerve zones as compared with noninjured control zones (bupivacaine P = 0.0003, lidocaine P < 0.0001, and ropivacaine P < 0.0001) and decrease in fiber area (bupivacaine P = 0.0002, lidocaine P = 0.0002, and ropivacaine P = 0.0002) denoting a drop-off of large-diameter fibers. There was also a trend toward smaller fiber density in lidocaine-injected (P = 0.0110) and ropivacaine-injected (P = 0.0102) groups relative to bupivacaine-injected nerves (P = 0.3433). No differences were observed between the drug-injected nerves across any measurements of severely injured areas (Fig. 4). No zone of severe injury was observed in saline-injected nerves as defined by the variables outlined earlier.
Axon–myelin ratios, previously described,35,36 were measured in all groups. The normal ratio is approximately 0.6, with a higher ratio indicating a decreased amount of myelin surrounding the axon. In the bupivacaine, lidocaine, and ropivacaine groups, the intermediate injury zones had an axon–myelin ratio of 0.68, 0.66, and 0.77, respectively. The severely injured zones had axon–myelin ratios of 0.76, 0.81, and 0.83, respectively. As indicated by the increasing ratios, there was a decrease in myelin thickness as injury progressed from the intermediate zone to the severe zone in all 3 experimental groups.
Cross-sections of the nerve were analyzed via EM and revealed that the extent and location of fascicular injury varied widely among the LA groups. In the saline group, no injury was observed apart from the occasional injury caused by the needle. Figure 5 represents an EM of an uninjured section of nerve in the saline group with normal-appearing myelinated and unmyelinated fibers. In the intermediate zone of injury, normal Wallerian degeneration was seen alongside smaller, uninjured fibers (Fig. 6). EM demonstrated decreased axon–myelin ratios along with some axons that maintained normal axon–myelin ratios. In the severe zone, EM revealed near complete loss of myelin along with increased fibrosis indicated by an increase in fibroblasts (Fig. 7). There was also increased collagen disorganization evident in this zone. Injury was also evident in the perineurium surrounding this zone, evident by perineurial thickening. In addition, edema and fibrosis were evident on EM throughout the endoneurium in all of our experimental groups.
Peripheral nerve block with LA is a common practice in providing pain control for a wide range of surgical procedures and pain syndromes. Inadvertent intrafascicular injection of an LA can generate a variety of nerve injuries, some of which may result in long-term disability.17 All anesthetics used in this study produced some degree of damage to the nerve when injected intrafascicularly, as evidenced by demyelination and Wallerian degeneration. The most severe injury occurred closest to the site of injection, and the extent of injury tapered off as distance from the injection site increased. This led to mixing of the injuries, evident in all LA groups on histological analysis as seen in Figure 4. Although there were minor differences in the extent of injury caused by the 3 LAs, there was an overall reduction of fiber quantity and fiber area in both the intermediate and severe injury zones. EM confirmed the severe injury zones by demonstrating absence of nerve fibers associated with severe fibrosis and collagen disarray. Intermediate injury was seen in all LA groups as areas of Wallerian degeneration alongside normal myelinated fibers. All 3 LAs used in this study—ropivacaine, lidocaine, and bupivacaine—caused considerable histological abnormalities when injected intrafascicularly.
Nerve injury from peripheral nerve blockade is well described; however, case reports of incidence, duration, and extent of sequelae are varied. Nerve injection injury is considered to be multifactorial in nature. Factors such as needle type and size, site of insertion, angle of insertion, pressure achieved during injection, type, and dose of medication injected (toxicity) affect degree of nerve injury according to various studies.3,6,9,10,12,18,37–39 All of these variables account for the disparity in the incidence of nerve injury with anesthetic injection reported in the literature.21,22,28,40
The effect of intrafascicular and extrafascicular injection of numerous drugs in common use has been extensively studied by our laboratory.2,3,5–7,9,10,12,38 In general, most drugs caused nerve injury when injected intrafascicularly, and, in contrast, extrafascicular injections produced little to no damage. A few exceptions, dexamethasone, botulinum toxin, and bovine collagen, demonstrated little to no axonal damage after intrafascicular injection.7,9,38 LAs have been shown to have a direct toxic effect on the nerve. Several studies have demonstrated that LAs can lead to fragmentation of DNA and disrupt the membrane potential in mitochondria, resulting in the uncoupling of oxidative phosphorylation. This in turn releases cytochrome c and activates caspase, which may result in apoptosis.41,42 The activation of proapoptotic enzymes in neuronal cells such as p38 mitogen-activated protein kinase and Jun N-terminal kinase have been shown to be initiated by lidocaine, bupivacaine, and ropivacaine.41,43 Generation of reactive oxygen species in Schwann cells by bupivacaine results in apoptosis.44 LAs have been shown to cause rapid necrosis of human neuronal cells. In addition, there is a direct correlation between concentration of the LA and time of exposure to the nerve with Schwann cell death.42,45 Rat, porcine, and canine sciatic nerves with exposure to LAs demonstrate infiltration of the nerve with macrophages and associated myelin damage.45 In addition to inducing apoptotic pathways and signals, LAs have been shown to constrict vasculature and decrease the blood flow to the nerves.46–48 There are a number of previous studies that have demonstrated differing levels of toxicity among various LAs.42,45,49 The observed toxicity from these previous studies is consistent with our study demonstrating that all 3 LAs produced neural damage.
In addition to toxicity of the medication used, the intraneural pressures achieved during accidental injection of drug into the nerve have been shown to be damaging. Hadzic et al.18 found that when using the same injectate volumes, intrafascicular injections resulted in higher pressures and were associated with lasting motor deficits and histological damage. More recently, Orebaugh et al.50 reported that all intrafascicular injections at the level of the brachial plexus root in fresh human cadavers result in high injection pressure. The saline control in this study demonstrated some areas of intermediate damage, comparable with that of the 3 anesthetic groups. Although we did not measure intraneural pressure, this may reinforce the idea that intermediate level injury can be largely attributed to the trauma of increased intraneural pressure on injection. However, the role of drug toxicity in producing intermediate level injury cannot be completely discounted as only 2 of the 4 nerves in the saline group showed intermediate injury whereas all 4 nerves in the bupivacaine, lidocaine, and ropivacaine groups showed intermediate injury.
In this study, intrafascicular injection resulted in varying grades of injury to the nerve as indicated by histomorphometry. Traditionally, peripheral nerve injury has been classified into 5 different degrees, as described by Sunderland.51 A first-degree injury, or neurapraxia, involves segmental demyelination. Second-degree injury, or axonotmesis, involves injury to both the axon and the myelin; however, the endoneurial tissue is not damaged. Third-degree injury involves an injury to the axon, myelin, and endoneurium and leaves the perineurium intact. A fourth-degree injury involves injury to the axon, myelin, endoneurium, and perineurium; the fascicle is scarred with only the epineurium being intact. Fifth-degree injuries occur when the fascicle or entire nerve has been transected. Sixth-degree injury was first described by Mackinnon et al.52,53 and involves a combination of first- through fifth-degree injuries. Fourth- through sixth-degree injuries require operative resection and repair. These types of injuries have been demonstrated by Sala-Blanch et al.38 where complete transection of fascicles (fifth- or sixth-degree injury) was observed. Our results demonstrate that all 3 LA groups demonstrated sixth-degree injury, as the nerves exhibited signs of first- to fifth-degree injuries. Saline-injected nerves (control) demonstrated areas of third-degree injury, with occasional axonal transection, or fifth-degree injury.
In summary, peripheral nerve injury from LA injection into the nerve occurs and remains a real clinical danger. The sequelae from such an injury may be long lasting and require surgical intervention. Intentional intraneural injection, as recommended by some clinicians to hasten the onset of the block, is not recommended.
Name: Scott J. Farber, MD.
Contribution: This author helped design and conduct the study, analyze the data, and write the manuscript.
Attestation: Scott J. Farber has seen the original study data, reviewed the analysis of the data, approved the final manuscript, and is the author responsible for archiving the study files.
Name: Maryam Saheb-Al-Zamani, MD, MS.
Contribution: This author helped analyze the data and write the manuscript.
Attestation: Maryam Saheb-Al-Zamani has seen the original study data, reviewed the analysis of the data, and approved the final manuscript.
Name: Lawrence Zieske.
Contribution: This author helped design the study and analyze the data.
Attestation: Lawrence Zieske has seen the original study data, reviewed the analysis of the data, and approved the final manuscript.
Name: Osvaldo Laurido-Soto.
Contribution: This author helped design the study and analyze the data.
Attestation: Osvaldo Laurido-Soto has seen the original study data, reviewed the analysis of the data, and approved the final manuscript.
Name: Amit Bery.
Contribution: This author helped design the study and analyze the data.
Attestation: Amit Bery has seen the original study data, reviewed the analysis of the data, and approved the final manuscript.
Name: Daniel Hunter, RA.
Contribution: This author helped design the study and analyze the data.
Attestation: Daniel Hunter has seen the original study data, reviewed the analysis of the data, and approved the final manuscript.
Name: Philip Johnson, PhD.
Contribution: This author helped design and conduct the study and write the manuscript.
Attestation: Philip Johnson has seen the original study data, reviewed the analysis of the data, and approved the final manuscript.
Name: Susan Mackinnon, MD.
Contribution: This author helped design the study and write the manuscript.
Attestation: Susan Mackinnon has seen the original study data and approved the final manuscript.
This manuscript was handled by: Terese T. Horlocker, MD.
1. Woolley EJ, Vandam LD. Neurological sequelae of brachial plexus nerve block. Ann Surg. 1959;149:53–60
2. Gentili F, Hudson AR, Hunter D, Kline DG. Nerve injection injury with local anesthetic agents: a light and electron microscopic, fluorescent microscopic, and horseradish peroxidase study. Neurosurgery. 1980;6:263–72
3. Whitlock EL, Brenner MJ, Fox IK, Moradzadeh A, Hunter DA, Mackinnon SE. Ropivacaine-induced peripheral nerve injection injury in the rodent model. Anesth Analg. 2010;111:214–20
4. Neal JM, Hebl JR, Gerancher JC, Hogan QH. Brachial plexus anesthesia: essentials of our current understanding. Reg Anesth Pain Med. 2002;27:402–28
5. Mackinnon SE, Hudson AR, Llamas F, Dellon AL, Kline DG, Hunter DA. Peripheral nerve injury by chymopapain injection. J Neurosurg. 1984;61:1–8
6. Strasberg JE, Atchabahian A, Strasberg SR, Watanabe O, Hunter DA, Mackinnon SE. Peripheral nerve injection injury with antiemetic agents. J Neurotrauma. 1999;16:99–107
7. Mackinnon SE, Hudson AR, Bojanowski V, Hunter DA, Maraghi E. Peripheral nerve injection injury with purified bovine collagen–an experimental model in the rat. Ann Plast Surg. 1985;14:428–36
8. Gentili F, Hudson AR, Kline D, Hunter D. Early changes following injection injury of peripheral nerves. Can J Surg. 1980;23:177–82
9. Mackinnon SE, Hudson AR, Gentili F, Kline DG, Hunter D. Peripheral nerve injection injury with steroid agents. Plast Reconstr Surg. 1982;69:482–90
10. Kobayashi J, Mackinnon SE, Langer JC, Hertl MC, Hunter DA, Tarasidis G. The effect of ammonium sulfate injection on peripheral nerve. J Reconstr Microsurg. 1997;13:389–96
11. Gentili F, Hudson AR, Hunter D. Clinical and experimental aspects of injection injuries of peripheral nerves. Can J Neurol Sci. 1980;7:143–51
12. Hertl MC, Hagberg PK, Hunter DA, Mackinnon SE, Langer JC. Intrafascicular injection of ammonium sulfate and bupivacaine in peripheral nerves of neonatal and juvenile rats. Reg Anesth Pain Med. 1998;23:152–8
13. Myers RR, Kalichman MW, Reisner LS, Powell HC. Neurotoxicity of local anesthetics: altered perineurial permeability, edema, and nerve fiber injury. Anesthesiology. 1986;64:29–35
14. Hadzic A Textbook of Regional Anesthesia and Acute Pain Management. 2007 New York, NY McGraw-Hill
15. Selander D, Brattsand R, Lundborg G, Nordborg C, Olsson Y. Local anesthetics: importance of mode of application, concentration and adrenaline for the appearance of nerve lesions. An experimental study of axonal degeneration and barrier damage after intrafascicular injection or topical application of bupivacaine (Marcain). Acta Anaesthesiol Scand. 1979;23:127–36
16. Fremling MA, Mackinnon SE. Injection injury to the median nerve. Ann Plast Surg. 1996;37:561–7
17. Auroy Y, Benhamou D, Bargues L, Ecoffey C, Falissard B, Mercier FJ, Bouaziz H, Samii K, Mercier F. Major complications of regional anesthesia in France: The SOS Regional Anesthesia Hotline Service. Anesthesiology. 2002;97:1274–80
18. Hadzic A, Dilberovic F, Shah S, Kulenovic A, Kapur E, Zaciragic A, Cosovic E, Vuckovic I, Divanovic KA, Mornjakovic Z, Thys DM, Santos AC. Combination of intraneural injection and high injection pressure leads to fascicular injury and neurologic deficits in dogs. Reg Anesth Pain Med. 2004;29:417–23
19. Clark WK. Surgery for injection injuries of peripheral nerves. Surg Clin North Am. 1972;52:1325–8
20. Villarejo FJ, Pascual AM. Injection injury of the sciatic nerve (370 cases). Childs Nerv Syst. 1993;9:229–32
21. Zhang J, Moore AE, Stringer MD. Iatrogenic upper limb nerve injuries: a systematic review. ANZ J Surg. 2011;81:227–36
22. Moore AE, Zhang J, Stringer MD. Iatrogenic nerve injury in a national no-fault compensation scheme: an observational cohort study. Int J Clin Pract. 2012;66:409–16
23. Iohom G, Lan GB, Diarra DP, Grignon Y, Kinirons BP, Girard F, Merle M, Granier G, Cahn V, Bouaziz H. Long-term evaluation of motor function following intraneural injection of ropivacaine using walking track analysis in rats. Br J Anaesth. 2005;94:524–9
24. Hara K, Sakura S, Yokokawa N, Tadenuma S. Incidence and effects of unintentional intraneural injection during ultrasound-guided subgluteal sciatic nerve block. Reg Anesth Pain Med. 2012;37:289–93
25. Robards C, Hadzic A, Somasundaram L, Iwata T, Gadsden J, Xu D, Sala-Blanch X. Intraneural injection with low-current stimulation during popliteal sciatic nerve block. Anesth Analg. 2009;109:673–7
26. Bigeleisen PE. Nerve puncture and apparent intraneural injection during ultrasound-guided axillary block does not invariably result in neurologic injury. Anesthesiology. 2006;105:779–83
27. Lupu CM, Kiehl T-R, Chan VWS, El-Beheiry H, Madden M, Brull R. Nerve expansion seen on ultrasound predicts histologic but not functional nerve injury after intraneural injection in pigs. Reg Anesth Pain Med. 2010;35:132–9
28. Barash P, Cullen BF, Stoelting R Clinical Anesthesia. 20096th ed Philadelphia, PA Lippincott Williams & Wilkins
29. Dunn P Clinical Anesthesia Procedures of the Massachusetts General Hospital. 2007 Philadelphia, PA Lippincott Williams & Wilkins
30. Klein SM, Greengrass RA, Steele SM, D’Ercole FJ, Speer KP, Gleason DH, DeLong ER, Warner DS. A comparison of 0.5% bupivacaine, 0.5% ropivacaine, and 0.75% ropivacaine for interscalene brachial plexus block. Anesth Analg. 1998;87:1316–9
31. Casati A, Fanelli G, Cappelleri G, Beccaria P, Magistris L, Borghi B, Torri G. A clinical comparison of ropivacaine 0.75%, ropivacaine 1% or bupivacaine 0.5% for interscalene brachial plexus anaesthesia. Eur J Anaesthesiol. 1999;16:784–9
32. Bertini L, Tagariello V, Mancini S, Ciaschi A, Posteraro CM, Di Benedetto P, Martini O. 0.75% and 0.5% ropivacaine for axillary brachial plexus block: a clinical comparison with 0.5% bupivacaine. Reg Anesth Pain Med. 1999;24:514–8
33. Kasukurthi R, Brenner MJ, Moore AM, Moradzadeh A, Ray WZ, Santosa KB, Mackinnon SE, Hunter DA. Transcardial perfusion versus immersion fixation for assessment of peripheral nerve regeneration. J Neurosci Methods. 2009;184:303–9
34. Hunter DA, Moradzadeh A, Whitlock EL, Brenner MJ, Myckatyn TM, Wei CH, Tung TH, Mackinnon SE. Binary imaging analysis for comprehensive quantitative histomorphometry of peripheral nerve. J Neurosci Methods. 2007;166:116–24
35. Mackinnon SE, Dellon AL, Hudson AR, Hunter DA. Chronic human nerve compression–a histological assessment. Neuropathol Appl Neurobiol. 1986;12:547–65
36. Mackinnon SE, Dellon AL, Hudson AR, Hunter DA. A primate model for chronic nerve compression. J Reconstr Microsurg. 1985;1:185–95
37. Jeng CL, Rosenblatt MA. Intraneural injections and regional anesthesia: the known and the unknown. Minerva Anestesiol. 2011;77:54–8
38. Sala-Blanch X, Ribalta T, Rivas E, Carrera A, Gaspa A, Reina MA, Hadzic A. Structural injury to the human sciatic nerve after intraneural needle insertion. Reg Anesth Pain Med. 2009;34:201–5
39. Selander D, Dhunér KG, Lundborg G. Peripheral nerve injury due to injection needles used for regional anesthesia. An experimental study of the acute effects of needle point trauma. Acta Anaesthesiol Scand. 1977;21:182–8
40. Liguori GA. Complications of regional anesthesia: nerve injury and peripheral neural blockade. J Neurosurg Anesthesiol. 2004;16:84–6
41. Lirk P, Haller I, Myers RR, Klimaschewski L, Kau YC, Hung YC, Gerner P. Mitigation of direct neurotoxic effects of lidocaine and amitriptyline by inhibition of p38 mitogen-activated protein kinase in vitro
and in vivo. Anesthesiology. 2006;104:1266–73
42. Perez-Castro R, Patel S, Garavito-Aguilar ZV, Rosenberg A, Recio-Pinto E, Zhang J, Blanck TJ, Xu F. Cytotoxicity of local anesthetics in human neuronal cells. Anesth Analg. 2009;108:997–1007
43. Lirk P, Haller I, Colvin HP, Lang L, Tomaselli B, Klimaschewski L, Gerner P. In vitro, inhibition of mitogen-activated protein kinase pathways protects against bupivacaine- and ropivacaine-induced neurotoxicity. Anesth Analg. 2008;106:1456–64
44. Park CJ, Park SA, Yoon TG, Lee SJ, Yum KW, Kim HJ. Bupivacaine induces apoptosis via ROS in the Schwann cell line. J Dent Res. 2005;84:852–7
45. Yang S, Abrahams MS, Hurn PD, Grafe MR, Kirsch JR. Local anesthetic Schwann cell toxicity is time and concentration dependent. Reg Anesth Pain Med. 2011;36:441–51
46. Kalichman MW, Myers RR. Transperineurial vessel constriction in an edematous neuropathy. J Neuropathol Exp Neurol. 1991;50:408–18
47. Kalichman MW, Lalonde AW. Experimental nerve ischemia and injury produced by cocaine and procaine. Brain Res. 1991;565:34–41
48. Kalichman MW. Physiologic mechanisms by which local anesthetics may cause injury to nerve and spinal cord. Reg Anesth. 1993;18:448–52
49. Radwan IA, Saito S, Goto F. The neurotoxicity of local anesthetics on growing neurons: a comparative study of lidocaine, bupivacaine, mepivacaine, and ropivacaine. Anesth Analg. 2002;94:319–24
50. Orebaugh SL, Mukalel JJ, Krediet AC, Weimer J, Filip P, McFadden K, Bigeleisen PE. Brachial plexus root injection in a human cadaver model: injectate distribution and effects on the neuraxis. Reg Anesth Pain Med. 2012;37:525–9
51. Sunderland S. The anatomy and physiology of nerve injury. Muscle Nerve. 1990;13:771–84
52. Mackinnon SE, Dellon AL Surgery of the Peripheral Nerve. 1988 New York, NY Thieme Medical Publishers
© 2013 International Anesthesia Research Society
53. Novak CB, Mackinnon SE. Evaluation of nerve injury and nerve compression in the upper quadrant. J Hand Ther. 2005;18:230–40