becker articol

7/29/2019 Becker Articol

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T he B io e lec tr ic F ac to rs in A m ph ib ian -L im bRegene ra t ion *

BY RO BERT 0 . BECK ER , M .D ., SYR ACU SE , N EW YO RKF rom th e D ep artm en t o f O rth opa ed ics, S ta te U niver sity o f N ew Y ork , U psta te M e4 uxi l C en ter ,

an d O rth opa edw S ect ion , V e terans Adm in istra tion H osp ita l, Sy rac use

T he rep lacem en t o f lo st bo dy pa rts by a spec ific g row th p ro cess k now n asregenera tio n is an ab ility sha red , to a v a ry in g ex ten t, by a ll liv in g th in gs. In gen-e ra l, a s o ne fo llow s th e ev o lu tio na ry sequen ce , the ab ility becom es in c rea sing lyrestr ic ted . In m an , the p rocess is lim ited to the reg en era tion of cer ta in tissueson ly , n o tab ly sk in an d bone . In a ch ild th e d iaph ysis o f a long bon e can be reg en -erated p ro v ided the pe rio steal tu be is in tact, w h ereas in an ad u lt a frac tu re o f alon g bon e can hea l b y rep lacem en t o f th e m is sing p ortion w ith fun ctio na l osseo ustissue . A ltho ugh the re is little d oub t th at fac to rs su ch as nu tr ition , vascu la r ity ,and im m obiliza tion all p lay im portan t ro le s in sup porting th is os seous hea lingprocess, the basic p roces ses o f ce llu la r de -d iffe ren tia tion and sub seq uen t re -d iffer-en t ia t ion w hich pro du ce a rep lica o f the m iss ing po rtion rep resen t true tissu eregenera t ion . T his p ro cess o f bon e hea ling is in m arked con tra st to the hea lingm echan ism s in m ost o the r hum an tissues in w h ich fib ro us co nn ec tive tissu e isused to b rid ge the gap . It is ev id en t, the re fo re, tha t th e p roces s o f reg en era tiveh ea ling m ust en tail spec ific facto rs no t op era tin g in o the r less adequ ate typ es o ftissue hea ling . To con tro l th is ty pe o f tissue healin g adequ ate ly , the se spec ificfacto rs m u st b e iden tif ied and them se lves con tro lled .

It is reaso nab le to expec t th at the sam e gene ra l co n tro lling m echan ism s areopera tiv e fo r a ll ty pes o f regen era tion , and tha t know led ge of the regene rativ epro ces s in ano ther v erteb rate m ay furn ish a b as is fo r a study of hum an regen era -t ion. For tuna te ly , the peak of reg ene ra tive ab ility , a s jud ged by the com plex ityo f the struc tu res reg en era ted , is p re sen t w ith in the ve rteb ra te phy lum . T he uro -d ela , o r sa lam ande rs , are p re sen t-day exam p les o f the basic ve rteb ra te ty pe fromw hich the h ig he r v erteb rate s have b een de riv ed . Th e ir fo re and h ind lim b s a reequ iva len t in ana tom ica l struc tu re and com p lex ity to the hum an upp er andlow er ex trem itie s . N eve rth ele ss , the la rva l-s tag e sa lam and er is capab le o f regen -e ratin g a p e rfec t rep lica o f on e of its lim bs in a pe riod o f th irty to fo rty d ay s. T headu lt sa lam and er is sim ila r ly cap ab le, alth ough the requ ired tim e is som ew ha tlonger and the regenerated extremity m ay be s lig h tly sm alle r than norm a l (F ig .1 ). T h is p rocess is so d ram a tica lly ev iden t tha t it ha s stim u la ted stud y fo r lite r-ally cen tu ries t. H ow ev er , the b asic con tro ffin g fac to rs in th e reg ene ra tive p roces san d the facto rs op e ra ting to red uce th e p rocess in m am m als h av e succes sfu llyre sis ted inv estig atio n u n til recen tly . W ith in the past few y ears , som e sign if ican tobserva tion s have been m ade and th e p rob lem now can be app ro ach ed from anew aspec t.

In on e lin e of in qu iry S inger d em onstra ted th e d ep en dence of the regene ra tivepro cess u pon th e p re sen ce o f a cr itic al am oun t o f ne rve tissue wi th in th e amputa -tion s t ump 23 , He showed that this effec t is no t depend en t on the type o f ne rv e

* Read at th e C om bined M ee ting of the O rthop aed ic Research Soc ie ty and Th e Amer i c anA cad em y of Or thopaedic Su rgeon s, M iam i B each , F lorida , Janu ary 8, 1961 .

t S p al l an z in i (Prodromo, 1768): But if t he above mentioned an im als recove r th eir legs -how com es it to pass, tha t o th er land an im als are n o t end ow ed w ith th e same powe r ? Is it t o behoped th at th ey m ay acqu ire th em by som e use fu l dispos i t ion?VOL . 43 -A . NO . 5 , JULY 1961 64 3


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64 4 R . 0 . BECK ERtissu e p re sen t. In the sa lam ande r , if the norm a l am oun t o f p e riph e ra l ne rv e tis sueis reduced by p rox im a l ne rv e sec tion in g , then regene ratio n w ill n o t occur un t i le ithe r the m iss ing ne rv es g row in to th e a rea o r un til neu ra l tissue o f an y type ,i)U t su ffic ien t in am oun t, is g ra fted in to the a rea . U nd e r ce rta in spec ific c ircum -stances, lim b regen era tio n can occu r iii l imb s d ev o id of n erves (parab io tic tw in -n ing 27 ) , in d icatin g tha t nu de r these c ircum stan ces the neural co n tr ibu tio n m aypo ssib ly be an cilla ry ra the r th an p rim ary . A ltho ugh these expe rim en ts hav e, ingene ral, lef t little dou b t th at som e neu ia l fac to r is necessa ry fo r the p ro cess o flim b regenera tion , the spec ific respo nsib le fac to rs assoc ia ted w ith th e nerv e haveye t to b e id en tified .

S om e recen t in vestiga tion s I rep orted fu rn ished a p ossib le c lue to the iden tityo f th is n eu ra l fac to r T he ex istence o f d irec t-cu rren t e lec tricity in liv ing organ -i sms w as first observed by G a lvan i in 17 92 , and has received a tten tion in te rm it-ten tly since then . T h is type of e lec tr ica l ac tiv ity , de sign a ted as th e bwelectr ic

F in . 1A n exam p le of the reg en erative ab ili ty of th e salam and er. Th e an im al o n th e lef t h :u l the r igh t

fo re lim b am puta ted on e w eek pr io r to the p ho tog rap h . T he other spec imen h ad the left fo relim brem ov ed at the sam e lev e l fifteen w eek s befo re. T h e regenera te is som ew hat sm a ller than the no r-m al lim b , bu t th e com pleteness o f resto ra tion is ev iden t. T hese fo rel im h s are h om olo gous to theh um an uppe r ex trem ity , con tain in g a hum e rus , rad ius , u lna, and carpal hones , iii add i t ion to th .eh om olo gous so ft structures. Th e reg ene rate res tore s alm o st comple te ly the ai ta to nucal com plex ityan d is a fun c tion al o rg an . It w ill be n oted th at the lo ng bon es, an d , pa rtic u la rly , th e sp ine of thespec im en w ith the w eek-o ld am puta tion , are less d is tin ct and app ea r som ewhat less den se than thesam e s tru ctu res o f the o the r an im a l. This o bse rva tio n has been m ad e repea ted ly and is no t du e toa ny d if fe re nc e in techniqu e. i ts s ign ific ance is unkn own a t l)re se nt.

THE JO UR NA L O F B O NE AN D JO IN T SUR G ER Y


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B IOE L E CTRIC FACTORS IN AMPH IB IAN -L IMB REGENERAT ION 64 5f ie ld or th e d irec t-curr en t fie ld , m ust be ca refu lly d istin gu ish ed from th e m orefam ilia r actio n -po ten tia l ac tiv ity o f the ne rves and m usc le s . T he b ioe lectr ic f ie ldis a steady-sta te p o ten tia l, m easu red as d irect-cu rren t po ten tia ls , on the su rfaceof liv ing organ ism s. The ac tion p o ten tia ls , on the o the r hand , co nsis t o f the trans-m ission of pu lses o f m em b rane depo la r iza tion a lon g the ne rv e o r m uscle f ibe r.A lth ough th e p u lse s are d etec ted by elec trica l m eans, th ey do no t inv o lve th eac tua l flow o f an e lec trica l cu rren t long itu d ina lly in th e fibe r S ince the tech -n iqu es o f m easu rem en t o f ac tion po ten tials are sim p le r , an d sin ce th e ac tion po ten -tia ls app a ren tly a re a p rim e m eans o f da ta transm is sion , th is typ e o f activ ityhas been pre fe ren tially stud ied b y m o st investiga to rs .

M uch of the w ork on the d irec t-cu rren t fie ld po ten tiaL s w as don e ten totw enty yea rs ago . T he concep t a rr ived a t th en w as tha t th e b io elec tric f ie ld con -sis ted o f a sim p le e lec tr ica l d ipo le w ith no m ore stru ctu ra l com p lex ity th an apo sitive po la rity o f the ceph alad po rtio n , com pared w ith th e caudad portion of thean imal . The source o f the po ten tia l w as assum ed to be the to ta l e lectr ica l activ ityof a ll o f the ce lls o f the o rg an ism . T he u sua l m easu rem en ts do ne a t tha t tim e w ereb etw een the n ose an d the ta il (ceph alad -cau dad) o r be tw een tw o o the r a rb itra ryp o in ts on ly . D esp ite th is am o rph ous concep t, m any ob se rva tion s w ere m ade ,in d icatin g som e re la tionsh ip be tw een these p o ten tia ls an d va rio us b io log ica lac tiv itie s such as: flu id tran sport in p lan ts 91#{176} ,anesth esia in an im a ls and ovu la -tion 19 , hypnosis 20 a nd sc hiz op hre nia 21 in m an . In pa rticu la r , th e re ap peared tobe correla tion s w ith g row th p rocesse s 6 , 15 , 16#{ 149} In recen t years the re la tionsh ip b e-tw een d irect cu rren ts and grow th w ere in vestiga ted by artific ia lly app ly ing thecurr en ts to the o rg an ism . A ccele ratio n of the ra te o f regene ra tiv e g row th in to -m ato p lan ts 23 and , con ve rse ly , com ple te reg res sion o f m a lignan t tum ors in m ice 12were repo rted to fo llow app lica tion o f cu rren ts o f ap pro priate po la r ity .

I t the re fo re app eared w o rthw h ile to re -ev alu ate the en tire p rob lem of th eb io elec tric fie ld an d its re la tionsh ip to g row th proces ses . T he sa lam ande r w asch osen as th e ex pe rim en tal an im a l in v iew of its regene ra tive ab ility and its pos i-tion in the v e rteb rate p hy lum . Furth e rm o re , th e m oist sk in o f th ese an im a ls p e r-m itted d irec t-cu rren t m easu rem en ts to be d one w ith le ss tech n ical d ifficu lty .U tiliz ing the tech n iq ues o f equ ipo ten tia l line p lo ttin g (de te rm in ing th e im ag ina ryline s on the su rface o f th e an im al w h ich co nnec t a ll p o in ts o f equa l po ten tia l) , acomple te fie ld p lo t o f th e b io elec tric fie ld can be m ad e. (F or fu rth er con cep ts o fthe e lec tr ica l f ield , the in te re sted reade r is re ferred to re fe ren ces 14 and 18 .). I fthe b io elec tric fie ld is a con tro lling fac to r in g row th pro cesses , such con tro l sho u ldb e in d ica ted by som e chang e from th e norm a l in the f ie ld p lo t. T h ere fo re , th ein itia l ob serva tion s w ere co ncen trated o n a de term in atio n of th e fie ld con figu ra -tion in the in tac t an im a l. R a th er than fin d ing the exp ected sim p le d ip o le , a com -p lex fie ld w as fo und w ith m u ltip le po sitive a rea s, o r elec tro n sink s, and m ultip lenega tiv e areas , o r elec tron sou rces (F ig . 2 ) . T h is p atte rn w as fou nd un ifo rm ly inove r fif ty in d iv id ua ls o f seve ra l d iffe ren t specie s . I t becam e appa ren t tha t thespa tia l con figura tion o f the b ioe lec tr ic fie ld co inc ided w ith the g ross ana tom icalarrangem ent o f th e cen tra l ne rvo us sy stem . T he positiv e a rea s (elec tro n s inks)co inc ided w ith the th ree m a jo r ce llu la r accum ula tion s o f th e n eurax is (c ran ial,b rach ia l, and lum bar en largem en ts ). T he nega tive a rea s (e lec tron so urces) co rre -lated w ith th e te rm ina tion s o f the m a jo r p eriph era l ne rve ou tflow s from thesesam e a reas. It w as the refo re po stu la ted tha t the d irect-cu rren t po ten tia ls w eregene ra ted w ith in th e ce llu la r aggrega tes an d d is tribu ted by th e pe riphe ral ne rves,w ith a com ple tion of th e c ircu it m ade th roug h th e so ft tis sues (o r in the aq ua ticfo rm s, th ro ugh th e en v ironm en t). T h is hyp o thesis w as tested in a num ber o f w ays.It w as n o ted th at com ple te sectio n (ad jacen t to th e co rd ) o f the n e rves to one ex-VOL . 43 -A . N O . 5 , JU LY 1961


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64 6 R . 0 . BE CKER

TH E JO UR NA L O F BONE A N D JO IN T S UR GER Y

trem ity p rod uced a p rom pt d rop o f th e po ten tial a lon g th e ex trem ity to nea rlyze ro . B y app ly ing th e H all e ffect (a phy sical m easu rem en t invo lv ing th e in te rac -tion be tw een a steady m agn e tic f ield and a m oving d irect cu rren t) it w as show nth a t th ere w as ac tua l f low of e lectr ical cha rges long itud in ally in an ex trem ity an dth a t th is f low ceased a fte r n erve sec tion . F ina lly , an ana logu e d ev ice, o r m od e l,o f th e cen tral ne rvou s system w as cons tru cted and w as foun d to p rodu ce th e sam efie ld p lo t an d to show th e sam e behav io r as the liv in g pro to type . It m ust be em -phasized , h ow ev e r, th at a ltho ugh these ex pe rim en ts show a d istinc t re la tionsh ipbe tw een the cen tra l ne rv ous system and th e b ioe lectr ic f ield , the spec ific tis suein the cen tral ne rvou s sy stem respon sib le fo r the g ene ra tion an d tran sm iss ion o fth e cu rren ts has no t b een id en tified . A t th is tim e w e can no t say w he the r the

FIG . 2Th e b ioe lec tnc fie ld , p rev iou s d ipo le concep t com pared w ith organ ized fie ld concep t as re la tedto th e c en tra l ne rvous sys tem .Th e ou tlin e draw ing on th e le ft ind ica tes th e tota l body po ten tia l as m ea su red by prev io us

inves tig ato rs (tip o f nose referen ce e lec trode , tip o f ta il reco rd ing e lec trode , an im al im m ersed influ id m ed ium ). T he equ ipo ten tia l lines as d raw n represen t the th eo re tica l d ipo le fie ld th at wa spostu la ted . S uch field d e te rm ina tions w ere never repor ted in the lite ra tu re . T h is is the type offie ld p lo t I expected to find w hen the pro jec t w as firs t begun .

T he m idd le ou tline d raw in g in d icate s the fie ld p lot that w as a ctu ally found (using th e tw o ele c-trod e eq u ipo ten tia l line -p lo tting techn ique on an an im al k ep t m o is t, bu t no t im m ersed , in so lu -tion ) . T he num erica l va lues inser ted a long the ex trem itie s a re typ ica l o nes found on an an im alabou t to recov er from an es thesia w h en the head s ink areas are u sed as the re fe ren ce e lec tro de siteand th e reco rd in g elec tro de is loca ted at the o th er appropria te s ites .

T h e ou tline draw ing a t the righ t rep resen ts th e g ross a rra ngem ents of the cen tra l n erv ou ssystem in the sa lam ander. It w ill be no ted th a t the neurax is con ta ins th ree m ajo r cellu la r accum u -la tio ns, w ith m ajo r periph eral ne rv e trunks ex tend ing d ista lly from each . T he co rresp ondencebe tween th e cel lu lar a ccum u la tion s an d the p ositive sink areas , an d be tw een the nerve-trun kterminat ions and the nega tive so urces, is apparen t .

neurons them se lves, the n eu ro p il, o r the su ppo rtin g e lem en ts-th e g lia an dS chw ann ce lls-a re th e im po rtan t struc tu re s .T hese obse rva tions on the b ioe lectr ic f ie ld in d icate th at it is a h igh ly organ ized

ac tiv ity , gene ra ted w ith in an d transm itted by elem en ts o f th e cen tral ne rv oussy stem . It appeared th a t, a s such , it cou ld func tion as a v ery p rim itive da ta trans-m ission an d con tro l sys tem . T h is co ncep t pe rm its a re study o f m any life p roces se s,inc lud ing grow th , w h ich a re ob v iou sly u nde r p rec ise co n tro l bu t fo r w h ich con tro lsy stem s h av e no t y e t b een id en tified . In th is p ap er I sh all be conce rn ed w ith lim bregen era tion , pe rh ap s the h igh es t ex p re ssio n of o rg an ized grow th in th e ad u lt


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B IO ELECTR IC FACTORS IN AM PH IB IAN -L IM B REG ENERATIO N 64 7

an im al. Th e d em ons tra ted re latio nsh ip b etw een the cen tra l ne rv ous system andthe b ioe lec tric f ield w ou ld ap pear to fu rn ish a lo g ical b asis fo r S in ge rs o bse rva -tion re la ting an y ty pe of n erve tis sue to reg ene ra tion . It ap pea red quite possib letha t the b asic p henom enon th at S ing e r observed w as n e rve -re la ted d irec t-cu rren telec tric ity ra the r than specif ic n eu ra l func tion . A pre lim in ary study of th is rela -tion sh ip be tw een the b ioe lec tric fie ld and grow th -co n tro l p ro cesses has neen com -p le ted and som e encou rag in g re su lts ob ta ined .

METHODS

Sev era l specie s o f salam and e rs w ith regene ra tive ab ility (T . v ir ide scen s,A . tig r inum , A . op acuum , an d N . m acu losu s) and a n on-regen era ting , bu t c lose lyre lated am ph ib ian , th e g ra ss frog (R . p ip ien s), w ere used as the exp erim en ta lan im als . Th e tech n iqu es o f m easu rem en t used w ere the sam e as p rev iou sly de -scr ibed 2 and w ill no t be d iscus sed he re . T he o n ly ch an ge in th is tech n iqu e hasb een a ref inem en t in the reco rd ing ap pa ra tus, p erm itting the sim ultaneo us m on i-to r ing of the d irec t-cu rren t vo ltages from as m any as e igh t d iffe ren t a rea s o f th es ame an im a l. In th e expe rim en ts in w hich d irec t cu rren t w as adm in iste red to th ean im als , a p rec is ion d irec t-cu rren t vo ltag e d iv ide r typ e of supp ly w as used a longw ith so lid , s ilv er-s ilve r ch lo rid e e lectrodes. O ne e lec trode w as large and en casedin a p ad soaked in sa line so lu tion to serve as th e refe rence e lec trode . Th e o th er w assm all and g la ss m oun ted and w as ap p lied d irec tly to the am pu tatio n stum p .A nim als w ere an es the tized w ith tricain e fo r m ost o f the exp erim en ts , alth ough thee ffec ts o f o the r anesth etic ag en ts on th e fie ld po ten tia ls w ere a lso stu d ied . D eepanesthesia , rega rd le ss o f the indu cin g agen t, is accom pan ied by a com ple te fie ldreve rsa l; th at is , th e ex trem ities becom e positiv e an d th e ce llu lar a rea s b ecom en eg a tiv e . D esp ite th is po lar ity rev ersa l, the equ ip o ten tia l line s rem a in th e sam eand th e fie ld p lo t rem ain s cons tan t. In te re stin g ly , the degree o f reve rsal o f f ie ldpo la rity co rre la te s w e ll w ith the dep th o f anesthesia , be ing fu lly rev ersed w ith th edeepest an es thesia an d slow ly re tu rn in g to no rm al as anesthesia d im in ish es .D uring th e p eriod (fiv e to ten m inu tes ) im m ed iate ly p reced in g fu ll reco ve ry fromanesthesia , the an im a ls d em ons tra ted a fie ld p lo t o f n o rm a l po la r ity and po ten -tia l. T h ese fac to rs w ere taken in to con side ratio n in m ak in g the m easu rem en ts .D ete rm ina tion s on w ound and reg en era te p o ten tia ls w ere don e just as th e an im a lsw ere fu lly recov ered from an es thesia. In som e instances, w h en re stra in t w as usedon non-an esthe tized an im a ls , s im ilar re su lts w ere ob ta ined .

RESULT ST he p resen ta tion of expe rim en ta l resu lts w ill b e d iv id ed in to fo u r sec tion s:

the e lectr ical ch anges im m ed ia te ly as soc ia ted w ith am pu ta tion ; the sub sequen tchanges during the h ea ling or reg ene ra tive phase ; th e e lec tr ica l chang es duringfrac tu re healin g ; an d th e resu lts o f a r tific ia lly augm en ting the elec trica l changesin th e co urse o f regene ra tio n .Sec tion O ne

L im b am pu tatio n in b o th reg en era ting an d non-reg en era ting fo rm s w as fo l-low ed im m edia te ly b y a com ple te reversal o f the e lectrica l po la rity a t the am pu-ta tion stum p (F ig . 3 ) . T h is change w as n o t a ssoc ia ted w ith any m a jo r chan ge inthe d irec t-cu rren t leve ls a t o th er site s m easu red , ex cep t th at the head vo ltagessub sequen tly sh ow ed con tinuo us flu ctu atio n (w av e fo rm s la stin g ten to f ifteensecond s) d u rin g the pe rio d of recove ry from an es thesia. These w ave fo rm s w erefo un d to b e assoc ia ted w ith an y m a jo r d am ag ing stim u li and , in th is ca se , theyprob ab ly rep re sen ted the con tin uou s stim u lus from th e am pu tatio n site . T hevoL . 43 -A . NO . 5 , JULY 1961


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64 8 R . 0. BE CKERrap id rev ersa l o f elec trica l po la rity a t th e site of am pu tatio n (w ith a sh ift fromnega tive to p ositive v o ltage ) m ay rep re sen t the cessatio n of d irec t-cu rren t dee-tron flow in th e asso cia ted n erve fibe rs . T h is ce ssa tion is a n eu ra l resp onse to th etraum a and does no t necess ita te sec tion ing of the nerve fibe rs them se lves. It w asno ted tha t any stim u lus p rodu cin g ce llu la r traum a-h eat co ag u la tion , rad iatio n ,freez ing , frac tu re o f th e lo ng bon e of an ex trem ity , o r crush in ju ry -p rodu ced asim ila r po lar ity sh ift, ob v iou sly w ithou t ne rve sec tion . T he du ra tion of the d irec t-

m VCHANNEL - - - - - NH E A D

+3 0 AMPUTAT IONLEFT FO RELEG-30

LEFT ________________FORELEG 0 ___________ ________

+3 0-30

BRACH IALAREA

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HD L EGC+3 0-30

R IGHT ________FORELEG

+3C0 10 20 30 40T IM E IN M INUTESFIG . 3

D ire ct-curren t vo lta ges m onito red at fiv e si tes b efo re and im m ediate ly a fte r am pu tation of thelef t fo re leg . D u ring deep anesthes ia (a t le ft on the tim e sca le ) a reversa l o f norm al field pa tte rn isp resen t, w ith the ex trem itie s p ositive and th e cen tra l a rea s nega tive to som e degree . A s anesthesiadec rea ses , the vo ltag es sw ing back th roug h zero to no rm al po la ritie s and po ten tia ls. A m puta tio nof the left fo releg in th is s tag e prod uces an im m ed ia te , la sting rev ersa l o f po la rity back to the sam elev els pre sen t d uring d eep an esthes ia . This po sitive v o lta ge n ot on ly pers ists to th e po in t o f fu llaw aken ing , b u t ac tua lly inc reases sligh tly in m agn itude . T h is p ositive vo ltage in th e le ft fo re lim b isac com panied b y a ten den cy to reve rsal o f po lar ity , re pre sen ted b y a slow e r rate of ch ang e, in thecon tra late ra l righ t fo re leg . T he le ft hin d lim b, how ev er, show s a S teady re tu rn to no rm al va lue .T he th ree lin es im m edia te ly abov e th e heavy lin e rep resen ting th e h ead -channe l vo ltage (top)refer to th e w av e-fo rm pattern s se en in th is chann e l w ith a fas te r sp eed of the record ing paper (o nem illim eter p er se con d) an d a h ighe r am p lific ation than w as used m the scale rep resen ted h ere. Indeep an esthes ia there is little o r no w av e-fo rm patte rn a t th is am p lifica tio n ; af ter amput a t i o n ,how ev er, con tin uou s w ave fo rm s app ear an d in cre ase in am plitude w ith tim e . T hese are the w av efo rm s ty p ical o f the response to traum a ; they a ctu ally represen t va ria t ion s in direct cu rren t sincethey have frequen cy per iods ranging from five to tw enty se co nds , In th is case , they ap paren tlyrep resen t th e respo nse to the co n tinu ous traum a aris in g from th e am pu ta tion .cu rren t respon se app ea red to be depend en t up on th e degree o f traum a . A t p resen t,w e can on ly specu late o n the m ech an ism re la ting the ce llu la r o r tissu e traum a tothe ch anges in th e b io e lec tric f ie ld itse lf. I t is appa ren t tha t th e traum a m ust besensed in som e w ay by the sys tem to bring abou t a ch ange in th e d irect-cu rren tac tiv ity o f the ap propria te cellu lar agg rega te (b rach ial o r lum bar). I t h as beenk now n fo r som e tim e th at traum a inv o lv in g ce llu la r d isru p tion causes a lo ca lcu rren t o f in ju ry 8 . P ossib ly , th is is the lo ca l m echan ism th at in f luences thed irec t-cu rren t ac tiv ity o f th e cen tra l n e rvo us system . Th at the re is som e re la-

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EXTREMITY

POTENTIAL

IN mV

5

F IG . 4

10 15 20DAYS FO LLOW ING

30 3 5 40

B IOE LE CTR IC FACTOR S IN AM PH IB IAN -L IM B REGENERAT ION 64 9

VOL . 43-A . NO . 5 , JULY 1961

t ion sh ip be tw een the pe rip he ra l ne rv e and the in ju ry po ten tia l is ev idenced byth e d ec reased curren t o f in ju ry e licited from ch ron ica lly dene rv ated stru ctu re s 28Sec tion Tw o

The sequ en ce o f chang es in th e b ioe lectr ic f ie ld d uring the h ea ling processe sin regen era tin g fo rm s w as v ery d iffe ren t from the fie ld -ch an ge seq uence in n on-reg ene ra ting an im a ls . In the fo rm er g rou p , a com plex se ries o f po ten tia l chang esoccurred tha t co rre la ted w e ll w ith the v ariou s stages o f th e regene ra tive p rocess(F ig . 4 ) . T h e in itia l p ositive p o la rity fe ll rap id ly in v alu e d uring the firs t tw o toth ree days, rose ag ain du ring th e fou rth to six th d ay s to a v alu e o f ab ou t 5 0 p er

25AMPUTAT ION

The vo ltag e chang es in an ex trem ity o f a sa lam ander during regenera tion afte r am putation . T heze ro p oin t on the h or izon ta l sca le imm ed iately precedes am pu tation , w h ich is con cu rren t w ith th every h ig h p ositiv e peak in th e vo lta ge curve. T he se con da ry po sit ive p eak in th is case is o n th e fifthday and the n ega tiv e m ax im um is on th e e igh th day . Th is n eg ativ e b ia s o f the regene ratin g l imb ism ain ta ined un til regen era tion is com plete . T he no rm allim b vo ltage is abou t ten m illivo lts nega tive(in terrup ted line ) . T h e low er f igu re s sh ow sch em a tica lly the stag es o f reg en era tion , w h ich in th iscase w as prac tica lly com ple ted by the th ir ty -eig h th to fo rtie th day .

cen t o f th e in itial, and then fe ll, cross ing th e ze ro lin e som e tim e be tw een the fif than d ten th day . T h is w as fo llow ed by a p rom pt n eg a tive sh ift o f h ig h am plitu de ,w ith m ax im um nega tiv ity o ccu rr ing som ew here b etw een the e igh th an d thefifteen th day . T h is nega tive po lar izatio n w as v e ry h igh , exceed ing in m illivo ltagebo th the no rm al lim b n eg ativ ity an d the p ost- traum atic p ositiv ity . Th is neg ativ ep o la rity rem ain ed e leva ted d uring the ph ase o f ax ia l g row th and d ifferen tia tion ,and as la te as the fo rtie th day a fter am pu ta tion the now a lm ost com ple te ly re-g en e ra ted ex trem ity w as still m o re nega tive than a ll the o the rs in m ore th an 50p er cen t o f th e an im a ls . T h is exp e rim en t invo lved a g ro up of fo u rteen an im a lsand , excep t fo r m ino r d iffe rences in th e am plitud e o f the v o ltages, they a ll fo l-low ed th e sam e sequ ence o f ch an ges.

The an a tom ica l and h is to log ica l changes du rin g the reg en e ra tive p rocess a red iv ided arb itra rily in to tw o phases. In itially , th ere is the accum ula tion , a t thes ite o f am pu ta tion , o f a m ass o f un d iffe ren tia ted cells . T he orig in o f these ce llsseem s to b e som ew hat in doub t, alth ough ce rtain quan tita tive m easu rem en ts have


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65 0 R . 0 . B ECKERrecen tly ind ica ted th a t th ey a re de rived from rem a in ing tis sue o f the lim b and tha tthe re is a m ov em en t o f ce lls from th e prox im al tissu es in to the stum p Th is u n-d iffe ren tiated cell m ass is k now n as th e b la stem a . It in creases in s ize w ith ou tev idence of d iffe ren tia tio n un til som etim e ab ou t tw en ty to tw en ty -f ive days a fteram pu ta tion (in the adu lt), w hen it beg ins to show ax ia l g row th and d iffe ren tiatio nand en ters the second phase . The o bse rv ed tim e co urse o f the vo ltag e -chang esequence sugg ests tha t th e in itia l p ositive p o la rity m ay b e assoc ia ted w ith re la-tive ly n on-spec ific healin g processe s o r w ith the b la stem a fo rm atio n , w he reas thenega tive po la r ity app ea rs to be de fin itely a sso c iated w ith the d ifferen tia tiongrow th ph ase. I t w as no ted tha t those spec ie s and ind iv idu als w h ich na tu rallyh av e a slow er rate o f regene ra tion show ed a co rre spo nd in g ly slow er ra te o f e lec -trical po ten tial change ; tha t is , the tw o processes w ere tim e re la ted .

EXTREM ITY10

POTENTIAL

IN m V o

NOR MAL0 - --

-20

- 2 4 6 8 1 0 12 14 16DAYS FOLLOW ING AMPUTAT ION

F IG . 5V oltage ch ang es du rin g the hea lin g process fo llow in g am pu ta tion of th e fo relim b of a frog (anon -regenera tin g fo rm ). T he zero po in t on the ho rizo n tal sca le im m ed ia te ly precedes am putation ,

wh ich is ag ain con curren t w ith the v ery h igh po sit ive po lar ity . U nlik e th e salam ande r (F ig . 4 )th is h igh pos itive po la rity is m ain tain ed fo r a lo ng per iod , dropp ing s low ly to nearly n orm al leve lsas hea lin g progresses a lm os t to com p le tion . T he no rm al fie ld pa tte rn in th is an im al is s im ila r to tha to f th e sa lam ander, an d th e ex trem itie s are in var iab ly negat ive du rin g the las t sta ge s of re cov eryfrom anesthesia (in ter rup ted line ).

A lth ough the no n-regene ra ting fo rm s dem onstrated the sam e im m edia tee lec tr ical-po lar ity rev ersa l fo llow ing am pu ta tion , they d id no t sh ow the sam e su b-seq uen t sequ en ce of e lec trica l-p o ten tia l ch an ges as th e regene ratin g fo rm s (F ig . 5 ).T he h igh ly po sitive p o ten tia ls rem ain ed e lev ated and fe ll on ly slow ly tow ard ze ropo ten tial a s h ea ling o ccurred . O n ly w hen hea ling w as com ple te d id th e ex trem itybeg in to sh ow neg a tive p o ten tia l and then it w as g en era lly low er than no rm al inva lue fo r som e days . I t m ust be emphasized tha t no t one of tw en ty -fou r sp ecim en sso exam ined show ed even a transien t sh if t to nega tive v o ltages u n til h ealin g w ascomp le t ed .

THE JO URNAL OF BONE AND JO INT SURGERY


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B IO ELECTR IC FA CTO RS IN A M PH IB IA N -L IM B REG ENER AT IO N 65 1T he hea ling p ro cess in these frog s con sisted in g ran u la tion tissu e and sca r

fo rm a tion w ith g radua l ep ith elia liz atio n o f the raw surface from the sk in edges(e ssen tially the sam e h ea ling proces s a s in m an ). T hese an im a ls a re rela ted to thesa lam ande rs , an d in th e la rv a l s tag es are them se lv es capab le o f com p lete lim bregene ratio n . Th ey lose th is ab ility in the adu lt fo rm . H ow ev er , th e adu lt fro gcan be m ade to regene ra te a nea rly com ple te lim b by gra f ting ex tra , func tion ingneu ra l tis sue in to th e am pu ta tion stum p 25 This is in te rp reted in the ligh t o fSingers o the r find ings as ind ica ting tha t the no rm a l density o f n e rve tissue inthe adu lt frog lim b is b elow the leve l necessary to susta in reg en era tion . E lec trica l

NORMAL

FRACTURE - -1J+IO

5 DAYS

10 DAYS 01F10

I 5 DAY S 0+10

FIG . 6Vo ltag e ch ang es o bse rved d uring frac tu re he aling . T he s im ple fractur e w as produced at th e

junct ion of the low er and m iddle th irds o f th e hum erus . Th e no rm a l v o lta ge g rad ien t progress ingpe rip he ra lly a long th e l imb is show n by th e u pp er- lin e graph . T his g rad ien t is measu r ed by p lac ingth e reference elec tro de ove r the b rach ia l are a an d sweep ing the reco rd ing elec tro de d ista lly . N o r-m ally , the re is a sm ooth cu rve o f inc reas ing nega tiv ity ob ta ined b y th is method .Th e low er fou r g raphs sh ow the d istu rb ances in th e lim b -vo lta ge grad ien t in duced b y th e fra ctu re

a t the tim es ind ica ted . T he seq uence of loca l v o ltag e changes is s im ila r to tha t show n by th e to ta lex tremity dur ing regenerat ion (F ig . 4) .

fie ld m easu rem en ts on in tact frog s dem onstrated a d irec t-cu rren t fie ld pa tternsim ilar to tha t o f th e salam and er , hav ing th e sam e spa tia l re latio nsh ip to th eana tom ica l a rran gem en t o f th e cen tra l ne rvou s system . S im ila r ly , traum a causedthe sam e po lar ity reve rsa l in the po ten tials . H ow eve r, the h ig h ly specif ic sequenceof po ten tia l chang es accom pany in g regen era tion in th e sa lam and er d id no t occu rin th e n on-reg en era ting frog . I have no t had the o ppo rtu n ity to m easu re th e p o ten -tia ls in a frog th a t w as regene ratin g a lim b secon da ry to ne rv e-g raf ting proced ures ,and a ttem pts to d rive th e elec trica l po ten tials o f the stum p artif ic ially in a regen-e ratin g sequ en ce fa iled because o f techn ica l p rob lem s rela ted to rep ea ted anes-thesia.VOL . 43 -A , N O . 5 , JULY 1961


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65 2 R . 0 . BECK ERSec tion Th ree

A fte r frac tu re o f a long bon e in the sa lam ander, the sam e po lar ity reversa lim m ed ia te ly ap peared w ith a sk in zon e of p ositiv e po ten tia l a ro und th e a rea o f th ebreak . T he subseq uen t vo ltage -ch ange seq uence du ring the h ea ling of the frac tu rew as sim ila r to tha t appea rin g du rin g lim b regene ratio n , a lthou gh som ew ha t low erin m agn itude (F ig . 6 ). T he d iffe rence in m agn itud e of the po ten tial w ou ld appea rto b e rela ted to the fact tha t the in ju red p a rt is w ith in the so ft tissu es and th at it iss itua ted a lon g the norm a l lim b vo ltage g rad ien t. T hu s fa r, no co rrela tion s haveb een m ade be tw een variou s s tag es of the f ractu re-h ealing pro cess and the vo ltag e-chan ge sequen ce , p r im arily b ecause o f the d iff icu lty in ob tain ing good roen tg en o-gram s of these sm all ex trem ities .

+3 I ; I CURRENT GROUP I

G IVEN 0 IN ju A

GROUP I fl-P pCONTROL n A -_ ---fl I? - iP

+3 GROUP 2CURRENT

G IVEN 0IN u A 23

PERCENT nGROWTH 0 IIINCREASE I I p

+3 GROUP 3CURRENT : CONSTANT CURRENT 2 iA /mm 2-

G IVEN 0 RANDOM OR IENTAT IO N OF AN IMALIN A .

-3 -20PERCENT 9

GROW TH 10INCREASE I & I I

00 5 0 5 20 25 30 35 40DAYS FOLLOW ING AMPUTAT IO N

F IG . 7The resu lts o f adm in istra tion of cu rren t o f various po la ritie s to the reg en era ting fo relim b of th e

salamander . The upper th ree lines (G ro up 1 ) re fe r to the se ries o f an im als g iven p ositive curren tdur ing th e fir st f ive day s af ter am puta tion . T he inc rea se in b la stem a form ation and the s low er ra teo f r e -d i ff e re n ti a ti o n in the exp er im en ta l an im als a re sch em atica lly show n. T h e cen te r tw o lin es(Group 2 ) re fer to the an im a ls g iven n ega tive cu rren t from th e fifth to the n in th day in clus ive(up per line ). T h e p ercen tage increase in ax ia l growth o f the ex perim en ta l lim bs ov e co n tro ls issh own on th e low er line . A p e riod o f g row th acce lera tion app ea red to ta ke p lac e f rom t he t we nt ie thto th e fo rtie th d ay , w ith a m ax im um accele ration abo ut the th irty -fif th d ay (23 per cen t). T helow er por tion (G roup 3) refers to the an im als ex po sed to con sta n t curren t , w ith ran dom orientat ionof th e an im al perm itted . T he so lita ry sp urt in ax ia l g row th o n th e tw en ty - fourth day is tho ugh t tohave been the resu lt o f changes in ion ic s treng th in the tank . In G roups 2 and 3 , a ll m easurem en tsw ere m ade w ith an ocu la r m ic rom ete r a t five to te n-d ay in terv als .

I t is an app a ren tly w e ll accep ted fac t th at frac tu re hea ling is a regene ra tiveprocess 17 This be ing the case , the observed regenera tive type of vo ltage-changeseq uence is en tirely app ro pria te . Th e re latio nsh ip b etw een th e density o f thepe riphe ral ne rves and the reg en era tive p rocess w as d ete rm ined on ly in the case

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B IO EL E CTRIC FACTOR S IN AM PH IB IAN -L IM B REG ENERAT ION 65 3of lim b reg en era tion . W ha t the rela tion sh ip is in th e case o f frac tu re hea ling is am a tte r o f specu la tion . A s prev iously sta ted , ne ithe r in ju ry to the ne rve itself no rp ro lifera tio n of th e n e rve fib ers need be con side red as p re req u isite s fo r the se -qu ence of e lectr ical-po ten tial ch anges. C e rta in ly , th e frac tu re is p rod uc tive o fsu ff icien t loca l cu rren ts o f in ju ry to in fo rm th e b ioe lectr ic f ie ld tha t su ch an even th as transp ired . Th e elec trica l-p o ten tia l ch an ges can be produ ced func tion ally bythe app rop ria te cellu lar agg rega te (b rach ial o r lum bar) w ithou t su ch ana tom ica lal terat ions.Sec tion Four

T hree sepa rate ex pe rim en ts w ere pe rfo rm ed in w h ich ex te rna l cu rren t w asadm in iste red to reg en era ting fo rm s afte r am pu tatio n . In th e firs t g rou p , s ix la rv alsa lam ande rs w ere g iven tw o m ic roam peres o f cu rren t d irectly to the am pu tatio nstum p fo r f iv e to ten m inu te s pe r day fo r th e firs t f ive posto pe ra tive d ay s. T h iscu rren t w as o rien ted so as to en hance th e norm a lly o ccu rrin g positiv e po lar ity a ttha t tim e. O n th e ten th to tw e lfth day , all s ix o f th ese an im a ls dem on stra ted ab la stem a a lm ost tw ice as la rg e as tha t no ted in a sim u ltan eo us co n tro l g roup . Itw as d ifficu lt to q uan tita te th ese d ifferences since the b la stem a is th ree-d im en -s io na l and so sm a ll tha t it does no t lend itse lf to p rec ise m easu rem en t. A n obse rva -tion o f eq ua l im po rtance to the inc rea sed size o f the b la stem a w as the su bsequ en tre tarda tion o f reg en era tion in the ph ase o f ax ia l g row th an d d iffe ren tiatio n . Ina ll o f the con tro l g ro up the re dev elo ped in itia l d ig ita tion of the reg en era te on th es ix teen th day , w he reas the exp erim en ta l g ro up d id no t reach th is stage un tilfo u r to s ix d ay s la ter . T hese resu lts a re sum m arized in F igu re 7 , u ppe r sectio n .

In th e secon d exp erim en t, s ix adu lt an im a ls w ere g iven th ree m icroam peres o fcu rren t d irec tly to the am pu ta tion s tum p fo r f ive to ten m inu te s pe r day from th efifth to th e ten th day . Th is cu rren t w as o rien ted to en hance the no rm a lly o ccu rr ingnega tive po la r iza tio n a t th at tim e . T he re w as no d isce rn ib le d iffe ren ce in b la stem afo rm a tio n com pared w ith a sim ultaneous con tro l g rou p ; h ow eve r , sub sequen t ax ia lg row th , a s m easu red by ocu la r m ic rom ete r a t in te rv a ls o f f ive to s ix days , ex -ceeded th at o f th e con tro l g roup by sig n ifican t am oun ts (F ig . 7 , cen te r sec tion ) .Th e com parison s o f g row th rate w ere m ade by m easu rin g by ocu la r m icrom e te rthe leng th o f the reg en era ted lim b in the six ex pe rim en tal an d s ix con tro l an im als .Th e ave rag e resu lts in each grou p w ere de te rm in ed , an d th e ave rag e pe rcen tag einc rea se in ax ial g row th o f th e expe rim en ta l an im a ls ov er th e co n tro ls w as cal-cu lated a t d iffe ren t tim e in te rva ls . It w as no ted tha t all o f the ex pe rim en ta l an i-m a ls show ed a greate r len g th o f reg en era te com pared w ith the m ost rap id ly g row -in g co n tro l an im a l. T he to tal s ize o f th e regene rate s , how eve r , w as sm all; a 25pe r cen t d iffe ren ce re fers to le ss than on e m illim e te r in ac tua l leng th .

D ig ita tion , a s an ind ex of d iffe ren tiatio n , also ap peared to be accele ratedin the exp erim en ta l g roup , bu t th is ob serva tion w as d ifficu lt to quan tita te . T h etech n ica l d ifficu ltie s encoun tered in th is exp erim en t necessita ted tha t sm a llnum bers o f an im a ls b e u tiliz ed . H ow ev er , d esp ite th e lack of a statis tic a lly sig n if i-can t sam ple , the co nsistency o f th e resu lts observed su ggest tha t som e grow th -acce le ra ting effec t w as ob tain ed . In th e an im a ls receiv ing p ositive p o la rity cu rren tfrom the firs t to the f ifth day a fte r am pu ta tion , an inc rea se in tissue res is tan cew as n o ted on the th ird day , n ecessita ting tw ice the p rev io us vo ltage to de live rthe sam e am oun t o f cu rren t. T h e sam e phenom enon w as n o ted on the e igh thd ay in the se rie s receiv ing nega tive po lar ity cu rren t from the fif th to the ten thd ay . In each case, cu rren t o f th e op posite po lar ity w as easily passed , w h ich w ou ldapp ea r to ind ica te tha t the re w as som e rec tif ier ty pe of m ech an ism in th e tis sues.T he sign ificance o f these obse rva tion s is n o t k now n .VOL . 4 3 -A , NO . 5 , JU LY 1961


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65 4 R . 0. BE CK ERA th ird g roup of six sa lam and ers w ere taken on the firs t d ay a fte r am pu ta tion

and p laced in a s tan da rd aqua rium w hich h ad a stead y d irec t elec trica l cu rren tf low ing th roug h the w a ter in o ne d irec tion . Th e cu rren t d ensity w as regu la ted top ro duce tw o m ic roam peres p er squa re m illim e te r, and th e an im als w ere pe rm ittedto assum e rand om orien ta tion in re sp ec t to th e d irectio n o f cu rren t f low . T he rew ere n o chang es in the ra te o f regen era tio n in th is g roup ex cep t fo r a so lita ry , bu tstrik in g , incr ease of ab ou t 19 p er cen t on the tw en ty -fo u rth to tw en ty -f ifth d ay ofm easu rem en t (F ig . 7 , low er sec tion ) . S ub seq uen t m easu rem en ts sh ow ed no s ig -n if ican t chang es in ra te, an d the g ro up com ple ted reg en e ra tion at abo u t th e sam etim e as the con tro l g ro up . In rev iew in g the exp e rim en ta l reco rds, it w as n o tedth at tw o days p rio r to the tw en ty -fou rth d ay the w a ter in the tan k w as su pp le -m en ted . S in ce K appe rs observed som e tim e ago tha t the trop h ic re spon ses o faqu a tic o rgan ism s to d irec t-cu rren t f ie ld s depen ded upon io n ic co ncen tra tion ,s im ila r fac to rs a re p resum ed to h av e been in ac tion h ere . T o ev alu ate p ro pe rlyth is random -o rien ta tion -s tead y-curren t s tudy , s trict con tro l o ver io n ic param -e ters w ou ld b e n ecessa ry .

S ince com p letin g th ese expe rim en ts , it ha s com e to m y a tten tion th at anex pe rim en t, s im ila r to th e la st on e descr ibed , w as don e w ith large n um bers o f an i-m als by F razee in 1909 . H e reported a sta tistically s ign ifican t inc rease in the ra teo f reg ene ra tion w hen d irec t cu rren t w as passed th rou gh tanks ho using la rv a lsalam ande rs w ith am pu ta ted lim bs. T he techn iq ues av ailab le at tha t tim e w erein su ffic ien t to ind ica te the cu rren t den sity , and know ledg e of the b io elec tricf ield w as fragm en ta ry . Th ese re su lts the re fo re cou ld no t th en b e in terp re ted inany sa tisfac to ry fram e of re ference . I t is o f in te re st to no te th at F razee had tracedsim ilar experim en ta l a ttem pts to in fluence the ra te o f regenera tio n by e lectrica lmea n s as fa r b ack as 1 840 22

DISCUSS IONG row th processe s are no t on ly com plex in na tu re bu t cove r a con tin uou s

spec trum of ab ilitie s an d po ten tia ls . W he the r one is cons ide rin g the em b ry on icg row th of an en tire o rg an ism from a single fe r tilized ovum o r the sim p le c losu reof a sk in de fec t b y ep ithe lial p ro lifera tion , the p rocess is qu ite ev iden tly un de rp rec ise con tro l . Tum or an d ke lo id fo rm a tio n a re usu a lly c ited as ev idence of losso r d istu rbance o f the con tro l m ech an ism s. W heth er o r no t th e sam e m ech an ism sare ope ra tiv e in a ll o f th e se co nd itio ns can on ly be specu la ted upon at th is tim e inv iew of the ex trem e pauc ity o f in fo rm atio n .

P h ilo so ph ica lly , the ph en om enon o f g row th is su ch a basic p rocess tha t o new ou ld expec t th at exceed ing ly p rim itive con tro l m echan ism s a re o pe ra tive an dth at a ll o f th ese p ro cesse s bea r som e in te rre latio nsh ip . Th e a fo rem en tioned w orko f S ing er , es tab lish ing th e re la tionsh ip b etw een th e pe rip he ra l n erves and lim bregene ra tion , is an im portan t b eg inn in g . H ow eve r, th is neu ra l effec t sho u ld no tb e in te rp re ted as invo lv ing any of th e know n fun c tio ns o f the n ervo us system .S in ce any ty pe of n erve tissu e is equ ally po ten t, such know n param ete rs a s th eac tion po ten tia l cann o t be in vo lv ed . It app ea rs m o re like ly th at som e v ery basicp ro pe rty o f the n ervo us sys tem , o r its a ssoc ia ted tissues, is the con tro llin g m ech-an ism . F rom th e ev idence pre sen ted in th is pape r , th e th esis is p ro posed th a t thed irec t-cu rren t ac tiv ity o f som e portion of th e cen tral ne rv ous sy stem cons titu te sa true da ta tran sm iss ion an d con tro l sy stem and tha t o ne o f the ou tp u t p a ram e-te rs o f the sys tem is the con tro l o f the g row th proces se s inv o lved in regen era tionan d rep air . S ince the e lec tr ica l-fie ld ac tiv ity is no t rela ted to the system tha tp ro du ced ac tion po ten tia ls and m ay even be a fun ctio n o f such tissu es a s th eS chw ann -ce ll sy ncy tium , the ob serva tion th at any ty pe of n erve tissu e w ill b r ing

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B IOE LE CT R IC FA CT OR S IN AM PH IB IA N -L IM B R EG EN ER AT IO N 65 5abou t regene ra tion b ecom es m ore log ica l. T h ere m ust ex ist cer ta in lin kage m ech -an ism s be tw een the ce lls o f th e o rgan ism and the b io e lec tr ic f ie ld itse lf, b o th in aninp u t o r sen so ry fa sh io n and as an ou tpu t o r con tro l m echan ism . C on side rin gsystem in pu ts , the re la tionsh ips dem onstra ted be tw een traum a and the chan gesin the f ield po ten tia ls app ea r to b e m ed ia ted by th e cu rren t o f in ju ry g ene ra tedat th e cellu lar lev el. R ega rd ing ou tpu ts o f the system , the in flu ence of a stead y-sta te d irec t-cu rren t p o ten tia l on ce lls h as b een prev ious ly dem onstra ted in thecen tral n ervo us system 26#{149}n th is ca se the rhy thm ic ac tiv ity (a s actio n po ten tials)o f co rtica l neu rons can be m odu la ted by chang ing th e d irect-cu rren t lev el ex ist-in g in the env iro nm en t o f the n eu ron s. M ore recen tly , it w as d em ons tra ted tha tce lls o f a reg enera ting tissu e h av e elec trica l ch arges th at are d iffe ren t from ce llsof the sam e tissu e in its n o rm a l sta te A n env ironm en ta l pa ram e te r fo r all ce llso f the o rg an ism is the d irec t-cu rren t o r b ioe lec tr ic fie ld po ten tia l g ene ra ted inth eir area. T h is f ield cou ld in f luence the s ite o f an in ju ry so as to cau se accum ula -tion of ce ll ty pes bea rin g sp ec if ic elec trica l cha rges a t po in ts hav ing sp ecif ic e lec -tr ic al po ten tials . In th is fash ion , the o u tpu t po rtion of the d irec t-cu rren t f ie ldsystem can b e lin ked to ce lls , and a true clo sed io op , d a ta tran sm iss ion and co n tro lsy s t em , is e stab lished fo r th e g row th p ro cess.

Th e p re lim ina ry na tu re o f these exp erim en ts m u st be s tre ssed . In all p ro b -ab ility , the e lec tr ica l f ie ld ch an ges accom pany ing grow th and hea ling processe sa re m uch m ore com p lica ted (bo th tem porally and spa tia lly ) th an are rep ortedhe re . N eve rthe le ss , it w ou ld appea r tha t ce rta in spec ific a ltera tions in the b io -e lec tr ic fie ld accom pany the p rocess o f regene ra tion and th at the se a lte ra tion s dono t app ea r in th e case o f le ss adequ ate h ealin g pro cesses . I t is su rp ris in g tha t theapp lica tion o f in term itten t e lectr ical cu rren ts p ro duced an acce le ra tion in thereg en era tive p rocess . If th e fie ld po ten tia ls a re the co n tro l m echan ism , theymus t be co nside red to be op era tiv e in a con tinuo us fa sh io n . In these expe rim en ts ,it m u st be assum ed tha t the observed a lte ra tion s w ere p ro duced in an an im a l w h ichno rm a lly has the capacity to regene ra te an ex trem ity , an d tha t th e added po ten -tials in th is case re in fo rced tho se w h ich w ere n atu ra lly o ccu rr ing . A lth ough itw ou ld app ea r th at con tin uou s cu rren ts w ou ld have to b e ap p lied to reve rse na tu -ra l po ten tia ls , the succes sfu l attem pt to inh ib it m alig nan t- tum o r g row th by in ter-m itten t f ield rev ersa ls m u st b e no ted 12

T he app lica tion of sim ila r tech n iqu es to the stu dy and con tro l o f g row thprocesse s in m an req u ires m uch m ore inv estig atio n bo th in an im als and in hum anbe ing s. O n ly recen tly h as a techn iq ue b een evo lv ed p e rm ittin g the p recise m eas-u rem en t o f th e d irec t-cu rren t f ie ld in th e h um an b ein g . In itia l m easu rem en ts m di-cated the p resence o f a h um an fie ld sy stem s im ila r to tha t o f the sa lam ande r, bea r-ing th e sam e spa tia l re la tion sh ips to th e cen tral ne rv ous sys tem . A nesth es ia w asfoun d to p rodu ce a sim ila r tend en cy tow ard reve rsal o f the f ie ld p o ten tia ls . H ow -eve r , the c ran ial po rtion s o f th e b ioe lec tric f ie ld in m an show a m uch greate r com -p lex ity than in an im als , appa ren tly be ing in fluenced by th e em otion s an d senso ryinp u ts. D esp ite th is , th e gen eral re la tio nsh ip be tw een th e hum an and the am phib -ia n b io elec tric f ield ar e rem arkab ly sim ila r.

In sum m ary , b ioe lec tric d irec t-cu rren t p o ten tia ls have been sh ow n to con -stitu te an o rgan ized d a ta transm ission and con tro l sy stem . R e latio nsh ips in d ica t-in g som e contro l fun ct ion have b een estab lished b e tw een th e d irec t-cu rren t fie ldan d the p ro cess o f lim b regen era tion .

REFERENCES1. BACKM AN , C . H ., an d B E C KE R , R . 0 .: T he Diffused Boundary E lec trod e . A S tab le M ethod fo rD . C . M easurem ents in the H um an (in prepa ra tion ).

VO L . 43-A , NO . 5 , JU LY 1961


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65 6 R . 0 . BECK ER2. BECKER , R . 0 .: T he B io elec tric F ield P attern in th e S alam ander and Its S im ula tio n by an

E lec tro n ic A na log ue . IR E T ran s. M ed . E lec tron ic s , 7: 202-207 , 1960 .3. BEN -OR , S. ; E ISENBERG , S .; and DouA N sxz, F .: E lec tro pho ret ic M obilities of N orm a l an d

Regene ra t ing L ive r C ells . N atu re, 188 : 120 0-1 201 , 19 60 .4. Ba& zm R , M . A . B .: T he E lec tric a l A ctiv ity of th e N ervous Sy stem . New Y ork , Th e M acm illa nCo. , 1960 .5. B UR GE , W . E . : F ur ther S tu dy on the E le ctr ica l T heory of An esth es ia . Am . J . Ph ys io l. , 1 26:

4 51 , 1 93 9.6. B URR H . S ., a nd HOVLAND , C . I. : B io-E le ctr ic C orrelates o f D eve lopm ent in Am bly stom a .Yale . B io l. and M ed ., 9 : 541 -549 , 19 37 .7. CHALKEY , D . T . : T he Ce llu lar B asis of L im b Regeneratio n . In Reg en erat ion in V e rteb rates,

pp . 34 -56 . Edited by C . S . Thorn ton . Ch icago , T he U niversity o f Chicag o P res s, 1 95 9 .8. CRAIB , W . H . : A S tud y o f the E lec tric al F ie ld S urroun din g Sk ele ta l M u scle . J. P hy sio l., 6 6 :

49-73 , 1928.9 . FEN SO M , D . S . : Th e B io-E le ctric Po ten t ia ls o f P lan ts an d Th eir Fu nct ion al S ign ific anc e.

I. A n E le ctro-K in etic Th eory o f T ransp or t. C anad ian J . Bo t., 3 5: 573 -58 2 , 1 95 7 .10 . FEN SO N , D . S . : T he B io -E lec tric P o ten tials o f P lan ts and Th eir F un ctio na l S ign ific anc e. I I. T heP atte rn s o f B io -E lec tric Po ten tia l and Exuda tio n R ate in E xc ised S unf low er R oo ts an d S tem s.C anad ian J. Bot., 3 6 : 367 -38 3 , 19 58 .11 . FRAZ E E , 0. E .: T he E ffe ct o f E lectr ica l S t im ulat ion upon the R ate of R egeneratio n in R ana

p ip iens and A m blys tom a je ffe rson ianum . J . E xp er . Z oo l., 7 : 457 -476 , 19 09 .12 . HUMP HR E Y , C . E ., and SE AL , E. H . : B iop hys ica l A pproach tow ard Tum o r R eg ress io n in M ice .Science, 130 : 388-3 90 , 19 59 .13 . KAPPERS , C . V . A . : F urther C ontrib u tions on N eurob iota x is . IX . An A ttem pt to C om pare the

Phenom en a of N euro b io tax is w ith O ther P henom ena of T ax is and T ro p ism . T he Dyn am icP o la riza tion of th e N euron e . J . C om p. N euro l., 27 : 26 1-29 8 , 191 7 .14 . KARPLUS , W . J. : Ana log Sim ula tio n . S o lu tion of F ield Pro b lem s . N ew Yo rk , M cG raw -H ill

B ook C o . I n c. , 1 9 58 .15 . KUR TZ , IRWIN , an d S C HR A NK . A . R . : B ioe lec tric a l P rop er ties of In tac t and R egenerat ingEarthw orm s, E isen ia Foe tid a. P hys io l. Z oo l., 2 8 : 322-3 30 , 1955 .16 . LU ND , E . J.: B ioelectric F ie lds an d G row th . A u stin , T he U niv ers ity P ublicat ion s, U n ive rsity

o f T exa s, 19 47 .17 . M CLEAN , F . C ., a nd URIST , M . R . : B on e. A n In trod uc tio n to the Physio log y o f Ske le ta l T is -sue . C hic ago , U n ive rsity of C hicago Press , 1 955 .18 . M ooN , P . H ., an d SPE NC E R , D . E . : F ounda tio ns of E le ctrodynam ic s. P rin ceton , N ew Je rsey ,V an Nostrand C o. Inc . , 1960 .

19 . PARMENTER , R ICHARD : D irec t Cu rren t P o ten tia ls A ssoc iate d w ith H um an O vu lation . Am . J.Physio l . , 126 : 59 7 , 1939 .

20 . RA vITZ , L . J . : E le ctrom etric C orrela tes o f th e H ypnot ic S ta te . Sc ien ce, 11 2 : 34 1-3 42 , 1 950 .21 . RAV ITZ , L . J . : E le ctrodynam ic F ield T heo ry in Psych iatry . South ern M ed . J ., 46 : 6 50 -66 0 ,

1953.22 . RU 5C ON I, M AU RO : Ueber KU nstliche B ebru ch tu ngen von F ischen and ilbe r e in ige neueV ersuche in B etref f k iln stlich er B efru ch tu ng an F r#{246} schen . A rch . A na t. Ph ysio l. u . W issen .Med., 185-193 , 1840 .23 . S imukh in , A . M .: B ioe lec tric Po ten tia ls in the R eg en erative Processes in P lan ts [title n R u s-s ian i. B io fiz ik a, 2 : 52 , 195 7 .

24 . SING ER , M AR CU S: Th e Influen ce o f the N erve in R egeneratio n o f the Am ph ib ian E xtrem ity .Quar t . Rev . B io l. , 2 7: 1 69 -2 00 , 1 95 2.25. S INGER , M AR C US : In du ct ion o f R egeneratio n of th e F ore lim b of the P ostm etam orp hic F ro g by

Augmentation of the Nerve Supp ly . J. Exper. Zoo!., 126: 419-471, 1954.26. T E R Z UOL O , C. A., and B UL L OC K , T. H.: Measurement of Impo sed Voltage G rad ien t A dequa te

to M odula te N eurona l F ir in g . P roc . N a t. A cad . Sc iences, 42 : 687 -694 , 195 6 .27 . YNT EMA , C . L .: R eg enerat ion in S pa rse ly Inn erv ated and A neurogen ic F orelim bs o f Am b ly-stom a L arvae . J . Exper. Z oo! ., 14 0: 101 -1 23 , 1 95 9 .28 . ZH IRM UN SKH , A . V .: O n the Parabio tic N a ture o f the R eaction o f M am m alian Sk ele ta l M u scleto D enerva tion [ title in R ussian i. F izio l. zh . S SSR , 44 : 577-585 , 1958 .

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