The Stomatogastric Nervous System of the Honey Bee

8/3/2019 The Stomatogastric Nervous System of the Honey Bee

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The Stomatogastric Nervous System of the Honey Bee(Apis mellifera ) in a Critical Phase of Caste Development

ISABEL C. BOLELI 1* , ZILA LUZ PAULINO SIMO ES 1 ,AND KLAUS HARTFELDER 21 Depto. Biologia, Facu lda de d e Filosoa, C iencias e Letras de R ibeiraoPreto, Universid ad e de Sao Pau lo, 14040-901 Ribeirao Preto, SP, Brazil2 Zoologisches In stitu t, U niv ersitat Tub ingen, D-72076 Tubi ngen, Germ an y*Correspond ence to: Isabel C. Boleli, Depto. Biologia, Faculda de deFilosoa Ci encias e Letras de R ibeirao Preto, US P, Av. B and eirantes 39 00,14040-901 Ribeirao Preto, S P, Brazil; E-m ail: zlpsim oe@usp .br

ABSTRACT Pr ogress in our u nder sta ndin g of polymorph ic differentia tion of female honey bee larvae into queens an d workers required a re-evaluat ion of neur onal pat hways potent ially involved in tra nsmitt ing informa tion on foodquality. This study presents new data on the anat omy of one of these path-ways, the stomatogastr ic nervous system (SNS) of honey bee larvae a ndpupae. Scanning electron microscopy preparations demonstrat ed not onlydevelopmental chan ges in frontal gan glion stru cture, but also provided rmevidence for a hypocerebra l gangl ion in the honey SNS. In addit ion topreviously described SNS nerves, the frontal , recurrent and esophagealnerves , and the fronta l connect ives , we observed three new nerves tha tconnect th e SNS to th e centr al nervous system a nd t he foregut. The rst one isan unpaired connective nerve of the frontal ganglion to the anteromedialprotocerebrum. The second consists of paired lateral branches of the recurrentnerve, and the third is a plexus of ne nervous branches associated with thepharynx. Lateral extensions of the newly described hypocerebral ganglionalso make contact with the pharynx. Similar but smaller branches were alsoobserved to or ig ina te from the esophageal nerves as they ru n a long theforegut . The exact an at omical localization of th e cardiostomat ogastr ic ner ves,which conn ect the SN S with t he r etr ocerebra l complex, could also be detected.The description of such n ew nervous connections will serve as a data base forfunctional analyses on the role of the SNS in differential feeding responses of the honey bee larvae, representing the initial step in caste differentiation. J.Morphol. 236:139-149, 1998. 1998 Wiley-Liss, I nc.

The polymorphic phenotypes observed inmost highly social insects do not result fromcaste-specic, distinct genetic program s, butrather represent hormonally controlled dif-ferences in morphogenetic growth. Th e rstand inductive step leading t o social insectcaste differentiat ion has been conceptual-ized as a nut ritiona l switch (Wheeler, 86).I n t he honey bee, Apis m ellifera , s u c h a

nutri t ional switch is well known and con-sists of the differential feeding of queen andworker larvae in the late larval stages witheither royal jelly or a mixed diet of glandu larsecre t ions , pol len and honey (Shuel andDixon, 59; Rembold et al., 74). Such exter -nal stimuli generally will require perceptionan d processing systems in order to tran smit

appr opriate informat ion to the endocrine sys-tem. The presence of chemical and mechani-cal receptors for food quality on mouthpartsof honey bee larvae has been demonstrated(Goewie, 78). Any further path ways in-volved in transmission and transformationof such information relevant to honey beecaste differentiation, however, still remainobscure, and only the nal element in su ch a

hypothetical a xis, the neu roendocrine t ract,has rece ived more a t ten t ion (Ulr ich and

Cont rac t g ran t sponsor : CAPES; Cont rac t g ran t sponsor :DAAD.

*Correspondence to: Isabel C. Boleli, Depto. Biologia, Facul-dade de Filosoa Ciencias e Letras de Ribeirao Preto, USP, Av.Bandeira ntes 3900, 14040-901 Ribeirao Pret o SP, Brazil.

JOURNAL OF MORPHOLOGY 236:139149 (1998)

1998 WILE Y-LISS, IN C.


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Rembold, 83). The ear liest cas te-specic en-docrine response is elevated levels of juve-nile hormone synthesis by th e corpora a llataof fourth and early-fth instar queen larva e(Rachinsky a nd Ha rt felder, 90).

The structure of the neuroendocrine tract(brain/corpora cardiaca/corpora allata) andits function in the regulat ion of synthesisand secret ion of morphogenetic hormoneswere e labora ted and are now wel l under-stood for a few lepidopteran species only, inparticular Mand uca sexta (Gilbert 89; West-brook et al., 91) an d Bombyx mori (Mizogu-chi et a l., 90).Att empt s to functionally char-acterize the neur oendocrine axis in the larvaland pupal sta ges of honey bee permitted usto identify specic sets of brain neur osecre-tory cells th at positively sta ined for a Bom-by x proth oracicotropic-hormone-like peptide

(Pa ulin o Simoes et al., 97). No immu nor eac-tivity a gainst this peptide, however, was de-tectable in a ny of the a xons. Similar indica-tions of an incompletely developed neuronalarchitecture in larval honey bees were alsoobtained for the serotonergic system (Boleliet a l., 95). Ser otonin, however h as been dem -onstrated t o stimulate juvenile hormone syn-th esis in vitro (Rachinsky, (94). These obser-vat ions led us to conclude tha t th e classicallyproposed neu roendocrine axis cont rolling in-sect metamorphosis may not be fully func-t iona l in the honey bee la rva , and tha t anadditiona l neuronal axis, the stomatogastricnervous system (SNS), could be involved in

transmitting information on food quality tothe endocrine system of the honey bee larva.

Turning to th e SNS a s a potential candi-date in the regulat ion of caste differentia-tion was encoura ged by th is systems wellestablished role in the coordina tion of foodintak e in insects an d food tra nsport th roughth e foregut (Hill et a l., 66; Dogra an d Ewen ,71; Verret and Mills, 76; Cooper and He,94; Miles an d Booker, 94). Connections of the SNS with the brain as well as with theendocrine complex have been reported inma ny in sects (Willey, 61; Gundel an d P enz-lin, 78; Kirby et al., 84; Dai et al., 87; Moorean d Loher, 88; J agota an d Habibulla, 92).

For th e honey bee, anat omical studies of theSNS date back over fty year s (Rehm, 40;Bickley, 42; Sch aller, 50; Rit cey a nd Dixon;69, Wirtz, 73). Despite all these accounts,however, the exact ana tomical organizationof the h oney bee SNS remains unclear andcontr adictory, an d man y deta ils concerninggut innervation, as well as connections of

the SNS wi th the bra in and the re t rocer-ebral endocrine complex, have not been re-solved. We, therefore, considered a detailedan alysis of ana tomical organization as a pre-

requisite for any further investigations intoth e SNSs ph ysiological role in cast e differen -t ia t ion . The purpose of th i s s tudy was tore-evaluat e th e gross m orphology of the lar-va l SNS in honey bee queen and workerlarva e by mea ns ofs cann ing electron micros-copy.

MATERIALS AND METHODS

Larvae and pupae of the honey bee (Apism ellifera L.) were obtained from colonieskept under n atu ral conditions at the Depart-men t of Genet ics, Un iversity of Sao P au lo atRibeirao P ret o, and th e Depa rt men t of Zool-ogy, Tubingen Un ivers ity, Germa ny. Queenlarvae and pupae were reared by graf t ingrst instar larvae as in standar d apiculturalpractice. Denition of substages in the criti-cal fth larval insta r wa s a ccording t o Rem-bold et al . (80) and Rachinsky a nd Hart-felder (90).

The SNS wa s exposed by an incision in t hecuticle on the back of the head, extendingthrough the neck region and thoracic seg-ments. Once opened, the preparation wasus h e d w it h p h os ph a t e -b u ffe r e d s a l in e(PBS), 0.1 M pH 7.4; NaCl 0.9%), and thetracheae and muscles surrounding the sto-ma togastr ic complex were carefully removed.Subsequ ent ly, PBS was replaced by cold xa-

tive (1% gluta raldehyde, 0.6% para forma lde-hyde, 0.25% DMSO in 0.12 M cacodylatebuffer at pH 7.2). The in si tu-xed brain,SNS, foregut, and retrocerebral complexwere then carefully dissected out of the headan d th oracic carcass, an d xed in fresh xa-tive for an additional 2 hr at 4C.

After r insing in cacodylate buffer, th epreparations were dehydrated in a gradedalcohol series up to absolute eth anol. Subse-quen tly, th ey were subm itted to critical pointdrying and, after spu tter-coating with palla-dium gold, they were examined and photo-graphed u sing Cambridge 250KM2 and J eolJSM 5200 scanning electron microscopes.

RESULTSThe frontal ganglion and its nerves

The s tomatogas t r ic nervous sys tem of honey bee female larvae and pupae is com-posed of two gangl ia , the f ronta l and thehypocerebral ganglion. The frontal ganglionis situated on the pharynx close to the fron-

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tal margin of the brain, between the phar-ynx e leva tor muscles (F ig. 1). I t i s pearsha ped, and its tip p oints ant eriorly (Fig. 2).When compar ing queens an d workers of thehoney bee, we noted n ot only cha racteristiccaste-specic differences but also chan ges inthe shape of the frontal ganglion during thelarval-pupal transition. In honey bee work-ers, the frontal ganglion has a typical pyri-form shape in t he la s t la rva l s t age , a ndbecomes a more rounded s t ruc ture in thepupa l phase, whereas t he fronta l ganglion of queen pupae is dorsoventrally compressed,giving it a n ellipsoidal a ppea ra nce (Fig. 3).

Five nerves exit the frontal ganglion: theunpaired frontal, recurrent and connectivenerves, and the paired frontal connectives(Figs. 16, 17). The frontal nerve originatesfrom the median anterior margin of the fron-

ta l ganglion . I t i s an unbranched ne andfr ag il e ne rve app rox ima te ly 1 0 0 m inlength, and runs forward along the dorsalmidline of the pharynx toward the labrum(Figs. 2, 3). The frontal connectives ar e a p-proximately 125 m in length and 15 m indiameter. They emerge from t he posterolat-eral m argins of the frontal ganglion, andt ravel to the an terovent ra l region of thet r i tocerebrum, jus t benea th the an tennallobes (Figs. 13). Before ent ering t he tr itoce-rebrum, each frontal connective merges withthe ipsilateral labral nerve to form a shortlabrofrontal nerve (Fig. 4). The labral nervei t se lf innerva tes the maxil la and labium.

The connective nerve arises from the medio-posterior margin of the frontal ganglion, jointly with the recurrent nerve (Figs. 5,16A). Over a short d ista nce, it rem ains fusedwith the recurent nerve before the ne con-nective nerve separa tes from th e dorsal sur-face of the stout recurrent nerve (Fig. 6). Itwas not possible to trace the complete pathof the connective nerve up to the posit ionwhere i t enters the protocerebrum becauseit is a very fragile structure that easily rup-tur es dur ing dissection.

The much stouter recurrent nerve is ap-proximat ely 45 m in diameter. It leaves theextreme posterior m argin of the fronta l gan-

glion between the two posterior pharyngealmuscles and travels posteriorly along themidline of the ph arynx (Figs. 3, 5) before itenters th e aorta an d nally reaches the hypo-cerebral ganglion. In al l specimens exam-ined, the recurrent nerve gives off two pairsof large lateral bran ches that a re symmetricin origin (Figs. 810). One of them forms a

peculiar ganglionic stru cture (Figs. 7, 9, ar-rows). In the same posit ion on the dorsalph a ry ngea l su r face, we observ ed m a n yperipheral neurons that are quite variablein th eir distr ibution patterns. The ne pro-cesses of these nerves a nas tomose an d forma complex nervous plexus that is l inked torecurrent nerve (Figs. 79, arrowheads). Inthe pos ter ior par t of the recur rent nerve ,where i t enters the anterior region of thehypocerebral ganglion, a pair of short andstout nerves emerges. These nerves corre-spond to the cardiostomatogastr ic nervesthat connect the SNS with the retrocerebralendocrine complex (Figs. 10, 11).

Th e hypocerebral ganglion an d its n erves

Being si tuated just beneath t he brain a nd

surr ounded by the retrocerebral system, thehypocerebral ganglion is not easily recog-nized. For clear demonst ra tion of th is impor-tant SNS element, i t was thus necessary tocarefully remove t he dorsal region of t hebrain and the retrocerebral complex in orderto expose and fully dissect this ganglion(Fig. 10). The hypocerebral ganglion of thepreimaginal honey bee then appears as anelongated and attened structure, closelyapposed to the dorsa l surface of the ph arynx.In d iameter i t i s s imi la r to the recur rentnerve (Figs. 11, 12).

Char acteristic for the larval h ypocerebralganglion are broad, n-like nerve branches

extending laterally over the pharynx (Figs.12, 13). The hypocerebral ganglion thus isnot only anatomically positioned closely overthe subretrocerebral foregut, but also ap-pears to be functionally connected to thisregion. Inter estin gly, this functiona l associa-t ion seems to be less in tense in the pupalphase, when width a nd density of such lat-eral extensions are much reduced (Fig. 11).

Two large nerves, generally described asesophageal nerves, arise from the posteriorregion of the hypocerebral ganglion (Figs.13, 14).After t ra veling a sh ort distan ce alongthe dorsal surface of the esophagus, thesenerves descend and run lateral ly on ei ther

side of the esophagus and crop (Fig. 14).Along its projection path, each esophagealnerve sends off numerous ten uous bran ches-that establish connections with the surfaceof th e poster ior foregut region (Fig. 13). Theesophageal nerves terminate in ve digit-like bra nches over t he lat eral su rface of thecrop (Fig. 15).

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Figures 16 Fig. 1. Apis mellifera. Frontal view of the br ain a nd frontal ganglion of worker pupae, showingthe local izat ion of the frontal ganglion between thephar ynx elevator m uscles (asterisk) . BR, brain; FG, fron-tal ganglion; LC, lateral connectives. Scale bar 50 m.

Fig. 2 . Apis m ellifera. Dorsal view of th e frontalganglion of a worker larvae. FG, frontal ganglion; LC,latera l connectives, Ph, pharynx. Scale bar 50 m.

Fig. 3 . Apis m ellifera. Dorsal view of th e frontalganglion of a queen pupae. FG, frontal ganglion; FN,frontal nerve; LC, lateral connectives; Ph, pharynx; RN,recurrent nerve. Scale bar 40 m.

Fig. 4. Apis mellifera. Lateral view of the frontal

ganglion. The frontal connective and ipsilateral labralnerve form the sh ort labrofronta l nerve before ent eringthe t r i tocerebrum. BR, brain; FC, f rontal connect ive;FG, frontal ganglion; IN, ipsilatera l labral nerve; LFN,labrofrontal nerve; Ph, phar ynx; PhM, pharynx elevatormuscle. Scale bar 50 m.

Fig. 5. Apis m ellifera. Front al ganglion removed fromthe pharynx to expose its ve nerves. CN, connectivenerve; FC, frontal connectives; FG, frontal ganglion; FN,frontal nerve; RN, recurrent nerve. Scale bar 50 m.

Fig. 6. Apis m ellifera. Detail of Figure 5 showing theconnective nerve emerging from the dorsal surface of therecurrent nerve. CN, connective nerve; FG, fronta l gan-glion; RN, recurrent nerve. Scale bar 10 m.


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Figures 711 Fig. 7. Apis mellifera. Dorsal view of the pharynx showing th e recurrent nerve, i ts nervousbranches ana stomosing in a ner ve plexus (arrowheads) ,and a peculiar ganglionic form at ion (arrows) . Oe, esopha-gus; RN, recurrent nerve. Scale bar 100 m.

Figs . 8 and 9. Apis mellifera. Detai ls of the la teralbranches of the recurrent nerve. Oe, esophagus; RN,recurrent nerve. Scale bar 10 m.

Fig. 10. Apis m ellifera. General view of the stomato-gastr ic nervous system showing the frontal ganglion,recurrent nerve, hypocerebral gan glion and esophageal

nerve. The brain was removed leaving only the basaltrit ocerebrum to support the prepara tion. CC-A, corporacardiacaaorta complex; FG, frontal ganglion; HCG, hy-pocerebral ganglion; OeN, esophageal ner ve; Ph, pha r-ynx; RN, recurr ent nerve. Scale bar 50 m.

Fig. 11. Apis m ellifera. Detail of the recurrent nerveshowing the cardiostomatogastric nerves emerging sym-metr ical ly a t the junct ion of the recurrent nerve withthe hypocerebral gangl ion. CC-A, corpora cardiaca-aorta complex; CSN, cardiostomatogastric n erve; HCG,hypocerebral ganglion; Oe, esophagus; RN, recurrentnerve. Scale bar 20 m.


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F i g u r e s

1 2

1 5


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Anatom y of the honey bee SN S

Schematic reconstructions of the SNS of preimaginal honey bees depict the a na tomi-cal position of the frontal ganglion and theprojection pattern of its nerves (Fig. 16A).Figure 16A shows in de ta il the yet unde-scribed connective nerve between the fron-tal ganglion and pars intercerebralis of theprotocerebrum. The posit ion of the newlydescribed hypocerebral gan glion an d th e in-nervat ion pa tter n of the crop by the esopha-geal nerves is reconstr ucted an d depicted inFigure 16B. A complete overview of the lar-val honey bee SNS (Fig. 17) shows the loca-tion of additional, newly described compo-nents of th i s nervous sys tem, such as thecardiostomatogastric ner ves connecting t herecurrent nerve to the retrocerebral endo-crine complex, as well as the nervous plexusformed by ne anas tomosing nerves tha texit the recurrent nerve as i t passes underthe tritocerebral bridge.

DISCUSSIONCom parative anatomy: newly d escribed

structures of the honey bee SNS

Scanning electron microscope an alysis of t he SNS o f honey bee l a rvae an d p upaeprovided a detailed view on the organizationof this nervous system. There is clear evi-dence for the existence of two ganglia, thea l ready well descr ibed fronta l gangl ion(Rehm, 40; Bickey,42; Ritcey an d Dixon,69), an d t he formerly obscure h ypocerebra l

gan glion. As th e hypocerebr al gan glion is a nimportan t str uctural element in the SNS of most insects (for review, see Penzlin, 85), its

absence in the honey bee, as postulated byRehm (40) an d r eported by Wirtz (73), waspuzzling. In hist ological cross s ections of th ehoney bee brain, the hypocerebral ganglionapparently could not be distinguished fromthe recurrent nerve as a separate enti ty. Aganglionic ma ss in this position, however,was suspected an d discussed by LHelias(50) and Schaller (56) as constit ut ing a tr uehypocerebra l ganglion . Our SEM guresshow tha t t his gan glion den itely exists. Itst ight associat ion with the foregut, and i tsposition close to the retrocerebral endocrinecomplex, however, ma y ha ve made t his st ru c-ture difficult to resolve in cross sections. Inother Hymenoptera, the hypocerebral gan-glion is a more dist inct s tructure (Lafon-Cazal and Verron, 80).

At the junction of the recurrent nerve withthe hypocerebral ganglion we could detect apa i r of n erves , the card ios tomatogas t r icnerves, which form a direct connection be-tween the SNS an d th e retrocerebral endo-crine system. This connection has alreadybeen noted by Ritcey a nd Dixon (69) andappears to be comparable to th e r ing n ervedescribed for the ant Lasius niger (Lafon-Cazal and Verron, 80). We have not yet beenable to completely trace the paths of thesenerves, and follow the projection patterns of their neu rons within t he endocrine system.

Another nerve tha t may be serving aninterface function between the SNS and theneur oendocrine a xis is the u npa ired connec-

tive nerve, a fragile nerve between th e fron-tal ganglion and the pars intercerebralis of the larval honey bee brain. The existence of such a nerve has not previous ly been re -ported for Apis mellifera , but was reportedin other insects, such as dragonies (Varma,72), crickets (Kirby et al., 84), cockroaches(Willey, 61; Gu ndel an d Pen zlin, 78), ter-mites (Hecker, 66) and scara b beetles (Bick-ley, ). In m ost insects, th e par s int ercerebra -l is of the protocerebrum is cons idered ahigher-order center that controls endocrineactivity. Neurosecretory material has beenlocated in the pars intercerebralis cells of honey bee la rvae by c lass ica l h i s tology

(Dogra et al., 77), an d m ore recent ly also byimmun ocytochemical localization of an in-sect prothoracicotropic h ormone-like pep-tide (Paulino Simoes et al., 97). Fur ther-m or e , a ca s t e-s p eci c t im in g for t h edevelopment of this neuroendocrine axis hasbeen reported (Ulrich and Rembold, (83).Based on our data, caste-specic differentia-

Fig. 12. Apis m ellifera. The hypocerebral gan glionof the larval SNS is a sa ddle-like stru cture connected tothe esophagus by short but broad la teral extensions.HCG, hypocerebral gangl ion; Oe, esophagus. Scaleba r 50 m.

Fig. 13. Apis m ellifera. Esophageal nerve leaving th ehypocerebral ganglion and projecting posteriorly alongth e esophagu s. HCG, hypocerebr al ganglion; Oe, esopha-gus; OeN, esophageal nerve. Scale bar 100 m.

Fig. 14. Apis mellifera. Esophageal nerves descend-ing on either side of the esophagus from a dorsal positionat their r oot in the h ypopharyngeal ganglion (top) . CC-A,corpora cardiacaaorta complex; Oe esophagus; OeN,esophageal nerve. Scale bar 100 m.

Fig. 15. Apis m ellifera. Finger-like termina l branch-ing of the esophageal n erve close to th e crop. Oe, esopha-gus; OeN, esophageal nerve. Scale bar 10 m.



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Fig. 16. Apis m ellifera. Schematic representat ion of the s tomatogastr ic nervous system of the larval a ndpupal honey bee. A: Frontal view. B : Hind view. AL,ant ennal lobe; Ao, aorta; BR, bra in; CA, corpora allata;CC A, corpora cardiaca-aorta complex; CN, connectivenerve; FC, frontal connective; FG, frontal ganglion; FN,

frontal nerve; HCG, hypocerebral ganglion; Mg, midgut;NCC, corpora cardiaca nerves; Oe, esophagus; OeN,esophageal nerve; Ph, pharynx; SOG, subesophagealganglion; 1st3rd TG, prothoracic, m esothoracic, a ndmetat horacic ganglia.


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t ion i s n ot l imi ted to the cent ra l nervoussystem but also can be detected in the fron-tal ganglion, and these caste-specic modi-cations may even, by their nervous connec-t ions to th e CNS, affect the activity of theprotocerebral center s of the n euroendocrineaxis during h oney bee meta morphosis.

The lateral branches of the recurrent n erveand the nervous plexus formed by the anas-tomosis of ne nervous processes of theperipheral neur ons observed on th e pharyn xare also an interesting and newly describedfeatu re of the honey bee SNS. Similar closeassociat ions between th e SNS and pharyn-geal surface were described in cockroaches(Willey, 61; Kha n, 76) an d in crickets (Kirbyet al., 84). An even tighter association be-tween SNS and foregut was apparent in thelateral extensions of the hypocerebral gan-glion of the larval honey bee. Such intr icateconnections have not been previously de-scribed in other insects, and their functionalrole is difficult to ascertain, since the honeybee SNS has received very little attention, sofar.

Fun ctional aspects

Detailed structural analyses of the larvaland early pupal SNS of the honey bee were

sought to provide a sound anatomical basisfor future functional studies, particularly onthe integration of food intake regulation andendocrine activity in caste differentiat ion.Functional analyses of the insect SNS havegenerally implicated t his system in the int e-gra tion of food inta ke a nd crop emptying (forreview see Penzlin, 85). Based on st ru ctura l

evidence, particularly the newly describednerve p lexus on the pharynx, the la tera lextensions of the hypopharynx, and the n-ger-like bra nching patt ern of the esopha gealnerves, we would expect this to be the casein the honey bee, as well . The interest ingquestion to ask in the honey bee, however,wil l be whether and how the SNS m ight beinvolved in cas te d ifferentia tion, e.g., by act-ing as a n int egrating relay between differentfood qualities (royal jelly vs. mixed workerdiet) an d th e endocrine system.

Functionally, the SNS is best chara cter-ized for locusts and cockroaches (for reviewsee Penzlin, 85). More recently, the sphingidmoth, Manduca sexta , h a s bee n ad de d t othis list because of its eminent m odel char ac-ter in hormonal control of metamorphosis.As demonstr ated for man y insects, th e fron-tal ganglion of M. sexta larvae turned out tobe a centra l pacemaker for n euronal activitycontroling buccal activity and muscle con-t r act ions i n t h e p h a ryn x an d e sophagus(Miles an d Booker, 94). Cas te-sp ecic differ-entiation of this ganglion in the honey beemay thus be conceived as a factor affectingfood int ak e rat es in honey bee caste differen -tiation.

Rhythmicity of foregut contra ctions, intur n, is expected t o be modulated by sensoryinput resulting from gut extension. A sourceof such sensory input could be the newlydescribed nerve plexus laterad of the recur-rent nerve. A similar plexus in locusts con-

sists of multipolar a nd bipolar neur ons (Fin-layson, 68; Kirby et al ., 84) The axonalte rminat ions of th ese neurons have beenencountered between the longitudinal andcircular muscles of the foregut and on thebasement membran e of the gut epithelium,suggesting that the peripheral nerve plexusmay be composed of both motor and sensoryne ur ons (Kirby et al., 84). Gelperin (67) an dMohl (72) demonst ra ted th at th ese m ultipo-lar neurons are stretch receptors, and fromelectrophysiological tra cing experiment s, Ta-bor (76) has s hown th at sensory neur ons of the pha rynx have branches both in th e recur-rent nerve an d th e fronta l connectives.

Whereas the SNS is t ra d it iona l ly per-ceived as a system regulat ing food intakeand gastrointest inal t ransport , i ts role as apotential relay between insect feeding activ-i ty and the endocrine system has recentlycome int o focus. When t ra cing a xons conn ect-ing the brain and retrocerebral complex inthe cricket Teleogryllus comm odus , Moore

Fig. 17. Apis m ellifera. Schematic general r epresen-tat ion of the s t omatogastr ic nervous system and i tsrelat ion to the foregut . CN, connect ive nerve; CSN,cardiostomatogastric nerve; F, foregut; FC, frontal con-nective; FG, frontal ganglion; FN, frontal nerve; HCG,hypocerebra l ganglion; NP, nervous plexus; OeN, esopha-geal nerve.



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an d Loher (88) discovered groups of somat ain the pa rs intercerebralis that project to theretrocerebral complex via the esophagealnerve. F urt herm ore, Reichwald et al. (94)

detected allatostatin-like peptides in nervebers of th e SNS of th e cockroach Diploptera punctata , an d Du ve et al. (95) were able toinhibit contraction of the foregut by treat-ment with callatostat ins. Serotonin, a modu-l a tory neu rot r ansmit t e r t ha t a lso s t im u-lates juvenile hormone synthesis in honeybee corpora allata (Rachinsky, 94), was notonly demonstrated in SNS neurons of manyinsects (Klemm et al., 86; Luffy a nd Dorn;91 Ha eften e t a l., 93), bu t wa s als o shown t oinh ibit gut cont ra ction (Ha eften et al., 93).The potential s ignicance in al latoregula-tion by s erotonin-immu noreactive cells inthe bridge connecting the corpora allata of

the larval honey bee (Boleli et al., 95) re-ceived further support by our nding thatthe cardiostomatogastric nerves conta ct theretrocerebral complex in this region.

To summa rize, the lar val and pu pal honeybee SNS is an atomically linked to the br ain,retrocerebral endocrine system, a nd foregutby several nerves and nervous connections.Axonal tracing and neuronal ma pping car-r ied out in other insect species disclosedcomplex neuronal pathways within the in-sect SNS and the systems i t is connectedwith (Gundel an d Penzlin, 78; Jagota andHabibulla, 92). Based on the anat omicalde ta il s of the la rva l honey bee SNS pre-sented in this study, i t should now be fea-sible to determine the role of this nervoussystem in the honey bee as well, part icularlyi ts role in the fascinating process of castedifferentiation.

ACKNOWLEDGMENTS

The authors are grateful to Prof. Dr. E.Laicine for providing access to the electronmicroscopy laborat ory of th e Fa culty of Medi-cine, Ribeirao Pr eto. Special t ha nk s a re du eto Horst Schoppmann, Zoological Institutein Tubin gen. Ma ria D.S. Fer reira an d J oseA. Mau lin, F aculty of Medicine of RibeiraoPr eto, for expert t echnical assisten ce in SEMprepar ations. CAPES an d DAAD na nciallysupported this study by a doctoral fel low-ship and a senior researcher fellowship.

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The Stomatogastric Nervous System of the Honey Bee