The facial nerve is often injured by trauma, infection or during the course of tumour resection. Many techniques of nerve anastomosis have been described with the current standard nerve repair using the microscope and monofilament suture. The purpose of this study was to evaluate the CO2 surgical laser as a tool for facial nerve anastomosis. Following preliminary electrical measurements 36 nerves were anastomosed using either laser or conventional monofilament suture. Laser anastomosis had neither beneficial nor detrimental effects on nerve regeneration. This method of anastomosis may be advantageous when surgical access is limited. In addition this study found that the use of CO2 laser as a dissecting or vapourizing tool in proximity to intact facial nerves results in degenerative change

Dort, Joseph C.

Felix, Heidi

Wolfensberger, Markus

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The Journal of Laryngology and OtologyJune 1994, Vol. 108, pp. 466-469CO2 laser repair of the facial nerve: an experimental study inthe ratJOSEPH C. DORT, M.D.*, MARKUS WOLFENSBERGER, M.D.f, HEIDI FELIX, P H . D 4AbstractThe facial nerve is often injured by trauma, infection or during the course of tumour resection. Many tech-niques of nerve anastomosis have been described with the current standard nerve repair using the microscopeand monofilament suture. The purpose of this study was to evaluate the CO, surgical laser as a tool for facialnerve anastomosis. Following preliminary electrical measurements 36 nerves were anastomosed using eitherlaser or conventional monofilament suture. Laser anastomosis had neither beneficial nor detrimental effects onnerve regeneration. This method of anastomosis may be advantageous when surgical access is limited. In addi-tion this study found that the use of CO, laser as a dissecting or vapourizing tool in proximity to intact facialnerves results in degenerative changes.Key words: Facial nerve; Laser surgeryIntroductionThe facial nerve is often injured by trauma or during thecourse of tumour resection. The sequelae of facial nerveinjury represent significant patient morbidity and there-fore improvements in nerve repair and regeneration aredesirable. Complete facial nerve injuries are currentlymanaged by means of micro-anastomosis with or withoutnerve graft interposition.Facial nerve recovery remains unpredictable withinconsistent clinical and experimental results (Wickholmet al., 1988; De Medinacelli and Seaber, 1989). Thisinconsistency may be explained by several local and bio-mechanical factors relating to the nerve repair but the mostimportant factor may be the random regrowth of axonsafter nerve transection (De Medinacelli and Seaber,1989). Recent investigations have led to the developmentof techniques that result in a more predictable recovery inexperimental animals (Wickholm et al, 1988; De Medi-nacelli and Seaber, 1989).The facial nerve presents a unique challenge to the sur-geon because of its complex course from the brain stem tothe facial musculature. The more distal mastoid and parotidportions of the nerve are easily amenable to suture repair.However, the labyrinthine and intracranial parts of thefacial nerve are more difficult regions in which to placesutures. Sutureless techniques have been used to make ana-stomoses in the labyrinthine and intracranial portions of thefacial nerve. However, the continuous pulsation of the cere-brospinal fluid (CSF) is a potential disruptive force to theanastomosis (Fisch and Mattox, 1988). In view of theseconcerns a method of reconnecting the facial nerve withoutthe use of sutures, such as CO, laser, is desirable.Experiments have shown that lasers can be used to'weld' tissue in vivo (Schober et al., 1986). The CO2 laserhas been used to join rat sciatic nerves without evidence ofharmful effects (Fischer et al., 1985; Beggs et al., 1986).Laser nerve anastomoses have been found to be weakerthan sutured anastomoses up until the fourth post-oper-ative day, after which both anastomoses were equallystrong (Maragh et al., 1988). Although several authorshave shown that the laser gives a superior neurographyresult (Fischer et al., 1985; de la Torre et al., 1988; Camp-ion et al., 1990) others have not observed a significantdifference between laser and suture repairs (Bailes et al.,1989; Benke et al, 1989).Much of the previous work has been done using the ratsciatic nerve model (Fischer et al., 1985; de la Torre et al.,1988; Wickholm et al, 1988; De Medinacelli and Seaber,1989). This model has been used extensively in peripheralnerve research and is stable and reliable. In addition, thesciatic function index (De Medinacelli et al, 1982) pro-vides a useful indicator of functional recovery after sciaticnerve repair. Nevertheless, it seems rational to use thefacial nerve as a model when one is studying the responseof that nerve to surgical manipulation. There has beenonly one study published investigating the use of the CO2laser in repairing the facial nerve and this was done in arabbit facial nerve model (Eppley et al, 1989). As pre-viously described there are several advantages to a ratfacial nerve model and therefore this has been used in thepresent study (Gleeson and Felix, 1986; Mattox and Felix,1987).This report examines the use of the CO2 laser for facialnerve anastomosis, comparing the results to suture ana-From the Department of Otolaryngology^: of the University of Zurich, Zurich, Switzerland, the Department of Otolaryngologyt, University ofBasel, Basel, Switzerland and the Department of Surgery*, University of Calgary, Calgary, Alberta, Canada.Presented at the Vllth International Symposium on the facial nerve, Cologne, June 9-14, 1992.Accepted for publication: 8 January 1994.466https:/www.cambridge.org/core/terms. https://doi.org/10.1017/S0022215100127124Downloaded from https:/www.cambridge.org/core. University of Basel Library, on 30 May 2017 at 18:54:10, subject to the Cambridge Core terms of use, available atCO2 LASER REPAIR OF THE FACIAL NERVE: AN EXPERIMENTAL STUDY IN THE RATE 467TABLE ICOMPOUND MUSCLE ACTION POTENTIALS! COMPARISON OF PRE-OPERATIVE AND POST-OPERATIVE RESULTS IN ALL ANIMALS ( ± SEM)Pre-operativePost-operativeCMAPAmplitude (mV)7.58 (0.64)3.95 (0.45)Latency (ms)1.15(0.03)1.59(0.10)stomosis. The report also evaluates the safety of using thelaser in proximity to the facial nerve.Materials and methodsExperiments were carried out on male Zbz SIV-50 ratsweighing an average of 250 gm. The animals were caredfor in accordance with the animal utilization guidelines ofthe University of Zurich. General anaesthesia was inducedwith ether and maintained with intraperitoneal pentobar-bital. Through a retroauricular incision the facial nervewas exposed while the buccinator muscle was exposedthrough a cheek incision (Gleeson and Felix, 1986). Con-stant current, square wave bipolar stimulation of supra-maximal intensity was applied directly to the exposedtrunk of the facial nerve. Evoked compound muscle actionpotentials (CMAP) were measured with silver needleelectrodes placed in the buccinator muscle and recordedon paper strips.Baseline electrophysiological recordings were made ina pilot group of four animals in order to ascertain the opti-mum recording conditions and normal values. These ani-mals were also used to assess the acute effects of the laserwhen used in proximity to the facial nerve. The nerveswere harvested one week after laser dissection, placed infixative and histologically examined.Eighteen animals (36 nerves) were placed into one offive experimental groups.Group I animals (n = 3: 6 nerves) underwent bilateralfacial nerve exposure with excision of a 5 mm segment ofnerve. This group functioned as a control and as a measureof the inherent regenerative ability of the rat facial nerve.Group II animals (n = 3: 6 nerves) had both facialnerves cut and then gently reapposed without sutures.In group III animals (n = 4: 8 nerves) the facial nerveswere cut and the epineurium was removed for a distance of3 mm on either side of the transection site. A single per-ineurial traction suture (10-0 nylon) was placed to facili-tate manipulation of the nerve. A series of six laser pulseswere then evenly spaced around the circumference of theanastomotic site. A microscope mounted CO2 laser(2W/50ms/defocussed/0.6 mm spot: Medilas AG, Dietli-kon, Switzerland) was used to make the anastomosis.In group IV (n = 4: 8 nerves) the facial nerves were cutand prepared as in group III. The nerves were then ana-stomosed by placing three 10-0 nylon perineural suturesequidistant around the periphery of the nerve.Group V (n = 4: 8 nerves) was identical to group IVexcept that the anatomosic site was also protected with asingle layer of Cargile® (Ethicon) membrane.At sacrifice (12 weeks post-operatively) facial nerveswere harvested for light microscopic evaluation. Thenerves were fixed in three per cent glutaraldehyde in 0. 1Mphosphate buffer for 24 hours. They were then placed inone per cent osmic acid in 0.1 M phosphate buffer for 120minutes and then dehydrated in graded concentrations ofethanol and embedded in Epon (Fisher ScientificCompany). Semi-thin sections were stained with toluidineblue and microscopically examined. Total counts of nervefibres were performed by hand on photomontages of thewhole nerve cross-section at a magnification of x540.Statistical evaluationThe results are presented as mean ± SEM. Compari-sons between groups were performed by analysis of vari-ance using the general linear model for unbalancedobservations followed where appropriate by Tukey'smultiple comparison test (Hassard, 1991).ResultsGeneral observationsObservations made one week post-operatively (in thepilot group) showed that there was some visible charringin the lasered nerves as well as a considerable perianasto-motic fibrous/inflammatory reaction in both sutured andlasered nerves. The charring resolved and the fibrous reac-tion was much less evident at 12 weeks. Group I animalsdid not demonstrate growth across the 5 mm nerve gap byeither electrical or histological criteria.Electrophysiological observationsElectrophysiological findings are presented in TablesI—II. Compound muscle action potential amplitude(CMAP) and latency are both adversely affected by sur-gery (Table I; P<0.001). However, intergroup compari-sons revealed no significant differences in CMAPregardless of the type of surgical repair performed. GroupI animals did not bridge the surgical gap and therefore noCMAP was measureable in this group. The Tables (I—III)are incomplete as several animals died in the peri-oper-ative period.Histology: qualitative observationsPerineural fibrosis and scarring was found irrespectiveof whether the laser or suture method was used to ana-stomose the facial nerve. Severe nerve reaction with totaldegeneration was found even when the laser was useddefocussed and at low power (2W/50ms/0.6 mm spot)within 1 mm of the epineurium. Morphology of the distalnerve fibres was abnormal in all groups. Myelin sheathswere thinner than normal and the intraneural organizationwas disrupted when compared to normal nerves (Figure 1 aand b).TABLE IICOMPOUND MUSCLE ACTION POTENTIALS (PRE-SACRIF1CE):INTERGROUP COMPARISONS ( ± SEM)GroupGroupGroupGroupII (« =I I I (n ••IV (nV(n =6)= 5)= 7)= 7)CMAPAmplitude (mV)3.35 (0.63)5.10(1.88)4.14(0.86)5.06(1.15)Latency (ms)2.00(0.10)1.71 (0.18)1.81 (0.06)1.69(0.07)n represents number of nerves.https:/www.cambridge.org/core/terms. https://doi.org/10.1017/S0022215100127124Downloaded from https:/www.cambridge.org/core. University of Basel Library, on 30 May 2017 at 18:54:10, subject to the Cambridge Core terms of use, available at468 J. C. DORT. M. WOLFENSBERGER. H. FELIXFIG. 1(a) Normal nerve fibres; (b) nerve fibres 12 weeks post-laser anastomosis. Note disruption and variability in myelin sheaths at this stage.(Toluidine blue; x540).Histology: quantitative observationsQuantitative nerve fibre data is shown in Table III.There was no significant different in fibre counts betweengroups. The distal/proximal ratio revealed increasednumbers of fibres distal to the anastomosis.DiscussionIn the evaluation of peripheral nerve regeneration fol-lowing traumatic injury it remains difficult to know whatparameters to measure. Return of function in the reinner-vated tissue is important and therefore the sciatic func-tional index is a clinically relevant measurement (DeMedinacelli etai, 1982). Electrophysiological and histo-logical data provide useful information about the func-tional and structural responses of injured nerves. Onedrawback to the rat facial nerve model is that there is novery good functional recovery scale. Subprimate mam-mals do not have the richness of facial innervation toallow meaningful interpretation of recovery by obser-vation alone.Eppley et al. (1989) compared suture and laser-assistednerve anastomosis in a rabbit facial nerve model. This isthe only published work comparing suture and laser ana-stomosis in a facial nerve model. These authors concludedthat laser anastomosis resulted in less entrapment of axonsat the repair site but unfortunately also produced thermaldestruction that could lead to a fibrous reaction. This studyshowed no electrophysiological differences between laserand suture anastomoses. A quantitative analysis of nervefibre regeneration was not performed in their study.The results of our study show that at the three-monthpost-operative evaluation both laser and suture repairscause very little grossly visible reaction at the anastomoticsite. Microscopically, both types of repair result in neu-TABLE IIIMEAN NERVE FIBRE COUNTS ( 1 O ' ± SEM)Proximal Distal Distal/proximalGroup II (n = 3)Group III (n = 5)Group IV (n = 5)Group V (/; = 3)6.71 (2.69)6.35 (0.82)6.86(1.04)4.71 (0.35)7.68 (2.67)8.28(1.56)11.44(0.79)10.81 (1.17)1.20(0.37)1.24(0.13)1.86(0.35)2.34 (0.36)n represents number of nerves.roma formation and extraneural escape of axons. Tensilestrength was difficult to assess in this experiment becausenone of the repairs were disrupted post-operatively prob-ably because of a tension-free anastomosis and an immo-bile wound.Compound buccinator muscle action potentials weremeasured in this study. The overall post-operative actionpotential amplitudes were significantly less than the pre-operative amplitudes reflecting incomplete and aberrantreinnervation of the buccinator muscle. Comparison ofaction potential amplitude between groups failed to revealany differences and therefore there are no electricallymeasureable differences in reinnervation regardless ofanastomotic technique. This conclusion has been reachedby others in different animal models. (Bailes et al., 1989;Benke et al., 1989).Nerve transection and repair causes major disruption ofnerve architecture which is not normalized even threemonths post-operatively. In addition to the late effects ofinjury, this study also found that using the laser in proxim-ity to the facial nerve resulted in significant nerve fibredegeneration, presumably secondary to the thermal injurycaused by the laser.Several studies have reported quantitative data on nervefibre regeneration after laser anastomosis (Fischer et al,1985; Beggs et al., 1986; de la Torre et al., 1988; Maraghetai, 1988; Bailes et al., 1989; Benke et al., 1989; Camp-ion et al., 1990) but none of the studies used a facial nervemodel. This information is usually reported in the form ofnerve fibre histograms. The hand counting method used inthis study, while more tedious, provides a more accuratehistological picture. The results show a trend to higherdistal fibre counts in the sutured nerves. However theseresults did not reach statistical significance perhaps due toinsufficient numbers of animals. If one assumes that anincreased number of distal fibres correlates with a betterquality of regenerative response then there is a trend tobetter regeneration in both the suture alone and the suture/Cargile® groups.ConclusionsThis study has shown that although the CO, laser is auseful technique for facial nerve anastomosis its use isneither harmful nor beneficial to nerve regeneration in thehttps:/www.cambridge.org/core/terms. https://doi.org/10.1017/S0022215100127124Downloaded from https:/www.cambridge.org/core. University of Basel Library, on 30 May 2017 at 18:54:10, subject to the Cambridge Core terms of use, available atCO2 LASER REPAIR OF THE FACIAL NERVE: AN EXPERIMENTAL STUDY IN THE RATE 469rat. The technique has the advantage of allowing a fasteranastomosis in a restricted surgical field. It is important toemphasize however that when used as a dissecting tool,even at low power settings, the CO, laser results in degen-eration of adjacent neural tissue. This finding may haveparticular relevance in the cerebellopontine angle andtemporal bone.AcknowledgementThe authors would like to thank Medilas AG, Dietlikon,Switzerland for lending the surgical laser used in theseexperiments.ReferencesBailes. J. E., Cozzens, J. W., Hudson, A. R., Kline, D. G., Ciric, I.,Gianaris, P., Bernstein, L. P., Hunter, D. (1989) Laser-assistednerve repair in primates. Journal of Neurosurgery 71: 266—272.Beggs, J. L., Fischer, D. W., Shetter, A. G. (1986) Comparativestudy of rat sciatic nerve microepineurial anastomoses made withcarbon dioxide laser and suture techniques. Part 2: A morpho-metric analysis of myelinated nerve fibres. Neurosurgery 18(3):266-269.Benke. T. A., Clark, J. W., Wisoff, P. J., Schneider, S., Balasubrama-niam, C , Hawkins, H. K., Laurent, J., Perling, I., Shehab, A.(1989) Comparative study of suture and laser-assisted anastom-oses in rat sciatic nerves. Lasers in Surgery and Medicine 9:602-615.Campion, E. R., Bynum , D. K., Powers, S. K. (1990) Repair of per-ipheral nerves with the argon laser: a functional and histologicalevaluation. Journal of Bone and Joint Surgery 72-A(5): 715-723.de la Torre, J. C , Karaca, M., Merali, Z., Fortin, T, Richard, M.(1988) Laser of razor?: a novel experimental peripheral nerverepair technique. Neurosurgery 22(3):531-539.De Medinacelli, L., Seaber, A. V. (1989) Experimental nerve recon-nection: importance of initial repair. Microsurgery 10:56—70.De Medinacelli, L., Freed, W. J., Wyatt, R. J. (1982) An index of thefunctional condition of rat sciatic nerve based on measurementsmade from walking tracks. Experimental Neurolog\ 11:634-643.Eppley, B. L., Kalenderian, E., Winkelmann, T., Delfino, J. J. (1989)Facial nerve graft repair: suture versus laser-assisted anastomosis.International Journal of Oral and Maxillofacial Surgery 10:50-54.Fisch, U., Mattox, D. (1988) Microsurgery of the Skull Base, GeorgThieme, Verlag, Stuttgart, pp 104-108"Fischer, D. W., Beggs, J. L., Kenshalo, D. L., Shetter, A. G. (1985)Comparative study of microepineural anastomoses with the use ofCO, laser and suture techniques in rat sciatic nerves. Part 1: Sur-gical technique, nerve action potentials and morphological stud-ies. Neurosurgery 17(2): 300-308.Gleeson, M. J., Felix, H. (1986) The Caletron ultrasonic surgicalaspirator system: an anatomical and physiological study of theeffect of its use on the rat facial nerve. Clinical Otolanngologv11: 177-187.Hassard, T. H. (1991) Understanding Biostatistics, Mosby YearBook Inc., New York, pp 88-89.Maragh, H., Hawn, R. S., Gould, J. D., Terzis, J. (1988) Is laser nerverepair comparable to microsuture coaptation? Journal of Recon-structive Microsurgery 4(3): 189-195.Mattox, D. E., Felix, H. (1987) Surgical anatomy of the rat facialnerve. American Journal of Otology 8: 43^17.Schober, R., Ulrich, F., Sander, T, Durselen, H., Hessel, S. (1986)Laser-induced alteration of collagen substructure allows micro-surgical tissue welding. Science 232: 1421-1422.Wickholm, R. P., Swett, J. E., Torigoe, Y, Blanks, R. H. I. (1988)Repair of severed peripheral nerve: a superior anatomic and func-tional recovery with a new 'reconnection' technique. Otolar-yngology-Head and Neck Surgery 99: 353-361.Address for correspondence:Joseph C. Dort, M.D.,Department of Surgery, Section of Otolaryngology,University of Calgary,Room 193A, Heritage Medical Research Building,3330 Hospital Drive N.W.,Calgary, Alberta,Canada T2N4N1.https:/www.cambridge.org/core/terms. https://doi.org/10.1017/S0022215100127124Downloaded from https:/www.cambridge.org/core. University of Basel Library, on 30 May 2017 at 18:54:10, subject to the Cambridge Core terms of use, available at

CO2 laser repair of the facial nerve: an experimental study in the rat

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