Table Of ContentReference: Bwl Bull. 180: 295-300. (April, 1991)
Control of Central and Peripheral Targets by a
Multifunctional Peptidergic Interneuron
DAVID PRIOR
J.
Department ofBiologicalSciences, Northern Arizona University, Flagstaff, Arizona 86011
Abstract. In the terrestrial slug. Limax maximus. feed- inson et al., 1988). alterations in feeding responsiveness
ingactivityandcardiovascularfunction havebeen shown (Prior, 1983; Phifer and Prior. 1985) and modifications
to be correlated. For example, in intact animals, both in cardiovascular function (Grega and Prior, 1986; Wels-
feeding responsiveness and heart activity are suppressed ford and Prior, 1991). Assuch, Umax representsa useful
during dehydration (Grega and Prior, 1986). The paired model for the analysis ofthe integration ofmultiple reg-
peptidergic buccal ganglion neurons RBI and LB1 have ulatory responses.
dramatic modulatory effects on both the feeding motor The concerted control offeeding behaviorand cardio-
program (FMP) and the force ofheart contraction (Wels- vascular function in Umax has been a focus of recent
ford and Prior, 1991). The Bl neurons appearto contain work (see Grega and Prior, 1985; Prior and Welsford,
the small cardioactive peptides (SCPs). Observations have 1989). Rhythmic feeding behavior in this organism in-
a frequency dependent excitation ofboth the FMP and volvesalternatingprotractionandretractionofthetoothed
the heart demonstrated by intracellular stimulation ofB1. radula against a food source. Feeding bouts often last
Thus, interneuron Bl mayserveto mediatethecoincident many minutes and can involve hundreds ofbite cycles
modulation ofmultiple responsestophysiological stresses. (seeGelperin etal.. 1978). Insemi-intactorisolatedprep-
arations of the central nervous system (CNS: Fig. 1),
Introduction chemical stimuli applied to the lips or electrical stimu-
lation ofthe lip nerves can elicit a prolonged pattern of
Environmental stress or a change in the physiological efferent neural activity that underlies the feeding move-
stateofan organism very often resultsin aconcerted array ments. Thisfeedingmotorprogram (FMP; PriorandGel-
of regulatory responses. Such responses usually include perin, 1977; Gelperin et al.. 1978) consists ofalternating
modification of behavioral patterns, or the level of be- burstsofactivityinprotractorandretractormotoneurons
havioral responsiveness,aswellaschangesin physiological (Fig. 1, 2). In addition to activation ofthe major buccal
functions such as cardiac output and respiratory activity. musculature, the FMP involves synchronized activation
With the use ofcertain invertebrate organisms, recent re- ofthe accessory salivary system. During feeding, the ac-
search has addressed the question ofthe control ofsuch tivity ofthe fast salivary burster neurons (FSBs), which
concerted response patterns (Prior, 1989; Teyke et al, arethe motoneuronstothesalivaryducts, becomesphase-
1990; Frugal and Brownell, 1987). locked with protraction (Fig. 2).
Terrestrial gastropods, such as Limax maximns, are
rdeehmyadrrkaatbiloyn.suIsnceaptdirbyliengtoenevnivriornomnemnetn,tatlhesytrecsasneslsosuech30a-s SCPB Modulation of Feeding and Heart Function
40% oftheirbody weight within a few hours (see Prior et In gastropods, the small cardioactive peptides (SCPs)
al., 1983; Riddle, 1983; Prior, 1985, for reviews). Among have an excitatory effect upon both the musculature (see
the array ofregulatory responsesdisplayed bydehydrating Lloyd and Willows, 1988; Lloyd, 1989) and the neural
slugs are contact-rehydration (Prior, 1984; Priorand Ug- networks underlying patterned feeding activity (see Wil-
lem, 1984), modifications in respiratory function (Dick- lows et al.. 1988). In several species, SCPB can initiate
CNS
patterned efferent activity in isolated preparations
Received 14 December 1990;accepted 30January 1991. (e.g.. Helisoma. Murphy et al.. 1985; Tritonia, Willows
295
296 D J PRIOR
.SN ci nl., 1988). In Limax, however, SCPB has a modulatory
role, increasing the responsiveness ofthe central pattern
BUCCAL
GANGLION generator to stimuli (Prior and Watson, 1987). In the
presence of 10 7 to 10 6 A/ SCPB, otherwise ineffective
stimuli can initiate full expression ofthe feeding motor
program.
Among the neurons in Limax that are responsive to
TENTACULAR o o SCPB are the paired fast salivary bursters. The rate ofen-
NERVE - dogenous burst activity in these motoneurons isenhanced
MCG MCG by application of SCPB in a concentration-dependent
CGAENRGELBIROANL manner(Fig. 3. 4). Short-term application ofSCPBresults
in a slow increase in FSB burst frequency and an even
ABDOMINAL
slowerdecrement ofthe effect follows initiation ofa saline
GANGLION
wash. In addition, continuous perfusion ofa preparation
for 20-30 min reveals no indication ofdesensitization of
M
the effect. In 10 6 SCPB. the burst frequency was sus-
tained at 14 bursts/min compared with a control fre-
quency of 1 burst/min (see Prior and Watson. 1987). It
hasbeen determined that thisexcitatory effect is mediated
by an increase in the rate ofthe interburst depolarization
BODY WALL rather than a general decrease in resting potential (Hess
Figure 1. A diagram ofthe isolated central nervous system ofLimax and Prior, 1989). Thus the effects ofSCPB on the Limax
including: the paired huccal gangliaand the fusedcerebral and abdominal feeding system include modulation ofthe responsiveness
ganglia;thepneumostome region;abdominal nervesN8-N12andthepos- of the FMP in addition to direct excitation of specific
teriorpedal nerves,PPN;buccal nervesN1-N3;gastricnerve,GN;salivary motoneurons.
SneBr;vec.erSeNb;robbuuccccaall pcroontnreaccttiovre,mCotBoCn;eumertoanccBr7e;brfaalstgsiaalnitvacrelyl.buMrsCtGer.neuron, Toassessfurtherthepotential roleofanSCPB-likepep-
tide in the regulation offeedingresponsiveness, exogenous
LSN I.I
*
I . , .
B
, lu| ,, ^u|
MULTIFUNCTIONAL PEPTIDERGIC INTERNEURON 297
UNI
I I I I I I I I
t
SCP ON
B
MINIMUM Ml M
2.
I I I I
SCP_ OFF
M
D
3.
I I I I I I ( I I I I I I I I I I I I.
B
LSB
IL
I
ll
T
SCP ON
D
Figure 3. (A) A continuous extracellular recording from the left salivary nerve (LSN) ofan isolated
buccal ganglia-brain preparation is shown in 1-3. The prominent bursting unit in this record is the faMst
salivary burster (FSB; each burst consists of 12-15 spikes). Within 20 s ofthe application of2 x 10~6
SCPBtothepreparation (first arrow), theburst frequencyofthe FSB increases. Followingremoval ofSCPB
from thesuperfusion medium(second arrow),burst frequencyoftheFSBreturnstothepretreatment level.
(B) an intracellular recording from the fast salivary burster neuron (FSB) showiMng the increase in burst
frequency and. in this case, progressive depolarization, in response to 2 I(T6 SCPB (the dashed line
indicatesthelevel oftheinterburst hyperpolarization beforeexposuretoSCPB). Bar = 30s(A)and 20mV
(B). (From Priorand Watson. 1988)
SCPB was injected into intact animals and their feeding and SCPA cause a concentration-dependent increase in
responsiveness measured. Asshown in Table I., SCPBcan the force ofventricular contraction (Welsford and Prior,
initiate the apetetive phases offeedingbehavior including: 1991). At a concentration of 10 6 M, SCPB can cause a
(1) cessation oflocomotion, (2) tentacular retraction, (3) 150% increase in the force of ventricular contractions.
lipeversion, and (4) lip movement. Thattheconsumatory Although lowerconcentrations ofSCPB ( 10~9to 10"7 M)
phase offeeding was not regularly initiated was not un- can cause a slight increase in heart rate, there does not
expected, in that in isolated CNS preparations SCPB did appeartobeaconsistenteffect(PriorandWelsford, 1989).
not initiate feeding, but rather, increased responsiveness Theexcitatory effectsofSCPBon heartand the feeding
to stimuli. Nevertheless, this would appear to be the first system ofLimax, together with the stress-induced coin-
demonstration ofan orderly effect ofinjected SCPB in an cidentchangesin feedingandcardiovascularfunctionob-
intact organism. This resultcertainly supportsthe notion served in intact animals (Grega and Prior, 1985), are in-
that an SCPB-like peptidergic system is involved in the dicative ofthe possibility ofcoincident control ofthese
control ofthe feeding system in Limax. two systems.
The small cardioactive peptides have been shown to
haveanexcitatoryeffectonthemusculature ofnumerous Multifunctional Modulatory Interneuron Bl
systems, includingHelixheart(Lloyd 1978, 1982),Aplysia
and Tritonia buccal mass and gut (Lloyd ct a/., 1984; In that exogenous SCPB can simultaneously modify
Lloyd and Willows, 1988), and Limaxventricle (Welsford feedingand cardiovascular function, immunohistochem-
and Prior, 1991; Lloyd, 1979; 1989). In Limax, both SCPB ical techniques were used in an effort to identify central
298 D. J. PRIOR
2XIO~*M
O O 2XIO"7M
A 2XIO"*
I.OXSALINE
Figure 5a. A photomicrograph ofa preparation of paired buccal
ganglia(outlined)showingthemorphologyoftheleftBl neuroninjected
with horseradish peroxidase. Cerebrobuccal connective. CBC; salivary
nerve. SN; thispreparation was madeby K. Delaney)
two major axonal projections and an extensive dendritic
A
arborization in the lateral lobe ofthe buccal ganglion.
smallaxonprojectsacrossthebuccal-buccalcommissure.
456
10
TIME(min)
Figure4. The responsesofthefastsalivarybursterneuron(FSB)to
vaningconcentrationsofSCPBarepresentedby plottingburst frequency
as a function oftime during the experiment. In each case, SCPB was
superfused overan isolated buccal ganglion-brain preparation between
minutes2and4.Thepreparationwassuperfusedwithsalinefor20min
between each trial. (A) The responses obtained in three trials with the
samepreparationusingvariousconcentrationsofSCPBareshown.Each 4
point representsthe burst frequency ofthe FSB in the preceding 60 s.
(B) The responses ofa second preparation to SCPB. In this case four
different concentrations ofSCPB were used as well as a control saline
trial.(C)Theextentofthevariabilitybetweenpreparationsisillustrated
by plottingthe mean (SD) burst frequency at each time point for29
trials in 12 preparations duringexposure to 2 x 10'6 A/SCPB. (From
Priorand Watson, 1987)
neurons containing SCPB-like-immunoreactive-material
(SLIM) that might be involved. Among the most prom-
inent SLIM-reactive neurons were the right and left Bl Figure 5b. Panel 1 isacamera lucidadrawingofthe somaofLB1
buccal neurons (Prior and Watson, 1987). In addition to following injection ofCo** showingthe majoraxon exitingthe buccal
those neuronsthatclearlycontain SCPB immunoreactive ganglionintheipsilateralcerebrobuccalconnective. Panel2isacamera
material,therearenumerouscellbodiesthatareenmeshed lucidadrawingoftheabdominalganglionshowingthecontinuedaxonal
bwyhicnehtiwsosrukgsgeosftiivemmoufnpoerpetaidcetrigviec efnidbeirnsgs(e.nge.a,rBt7h,e tFaSrBg)e,t gapranontjgieldcirtooinmoinnceorafvcettih9veaatinindojnetcwotofedBalxLoBni1na,lrwebisrtpahonncsoheneestoianrxenopeneratvlietibv1re1a.nsPctaihnmeuillna3atibioldlnoumsotifrnattaheles
feeding neurons. cardiac branch ofnerve9. Panel4shows repetitiveintracellularstimu-
The morphology ofBl was examined by intracellular lation ofBl causing in a constant-latency axonal impulse recorded in
injection of horseradish peroxidase (Fig. 5a). There are nerve9.
MULTIFUNCTIONAL PEPTIDERGIC INTERNEURON 299
lahlc I
BehavioraleffectsiifSCPg injeclums in Limax maximus
Behavioral observations
Treatment
300 D. J. PRIOR
salivary bursters. As shown in Figure 6, driving Bl at 5 Hess,S.D.,and D.J.Prior. 1989. SmallcardioactivepeptideBmod-
impulses/s can result in a 50% increase in FSB burst fre- ulates feeding motoneurons in Limax maximus J Exp. Biol. 142:
473-478.
quency. Even two to three impulses at low frequencies Lloyd, P. E. 1978. Distribution and molecular characteristics ot car-
are sufficient to elicit a transient increase in FSB burst dioactivepeptidesinthesnail.Helixaspersa.J. Comp. Physiol. 128:
frequency. These effectsare sustained in high Mg++. high 269-276.
Ca++ saline indicating the possibility of monosynaptic Lloyd, P. E. 1979. Central peptide containing neurons modulate gut
connection. activity in Trilona. Soc. Neurosci. Abstr. 5: 252.
Toassessthepotential roleofBl in thecontrolofheart Lloy4d1,:P2.94E.8-12988522.. Cardioactiveneuropeptidesingastropods.Fed. Proc.
function, semi-intact preparations ofthe CNS and inner- Lloyd, P.E. 1989. PeripheralactionsoftheSCPsinAplysiaandother
vated heart were used, which allowed intracellular stim- gastropod molluscs. Am. /.ool. 29: 1265-1274.
ulation of Bl and measurement of ventricular activity. Lloyd, P. E., I. Kiiplrim.urn, and K. R. Weiss. 1984. Evidence for
Stimulation of Bl at low frequencies resulted in an in- parallel actions ofa molluscan neuropeptide (SCPA) and serotonin
inmediatingarousalinAplysia Proc.Nat.Acad.Sci. L'SA81:2934-
crease in the force ofcontraction ofthe heart (Fig. 7). It 2937.
isofinterest that Bl frequencies of5 to 7 impulse/s were Lloyd,P.E.,andA.O.D.Willows. 1988. Multipletransmitterneurons
the most effective in the activation ofboth the FSB and in TritimiaII.Control ofgut motility. / \eurohml. 19(1): 55-67.
the heart. Murphy, A. D., K. Lukowiak, and \V. K. Stell. 1985. Peptide modu-
batWhheedninthhiisghexMpegrfifm,enhtighwaCsar+e+pesaatliende,witthehrethweasCNnSo PhiflPearrt,oicoCn.NoBaf.t,.paaAtntcdearDdn.eJdS.cim.PortiUooSrr.Aa1c89t28i:5vi.7ty14iB0no-i7dd1ye4nh4tiy.fdireadtinoenuraonndshoafeHmcloilsyommpah.
changeintheeffectivenessofBl toincreaseheartfunction. osmolality affect feeding and its neural correlate in the terrestrial
This suggests that B1 hasa direct effect on peripheral tar- gastropod, Limaxmaximus. J. Exp. Biol. 118: 405-426.
getsratherthan actingviaadditional CNS neuronal path- Prior, D. J. 1983. Hydration-induced modulation offeeding respon-
sivenessin terrestrial slugs.J. Exp Zool. 227: 15-22.
ways. Prior, D.J. 1984. Analysisofcontact-rehydrationinterrestrialgastro-
Thus, it would appear that buccal neuron Bl may be pods: osmoticcontrol ofdrinkingbehaviour. J Exp. Biol 111:63-
a multifunction peptidergic interneuron capable of si- 73.
multaneously modulating the central feeding motor pro- Prior, D.J. 1985. Water-regulatory behaviourin terrestrial gastropods.
Biol. Rer. 60: 403-424.
gramandcardiovascularfunction. Assuch, Bl, alongwith
Prior, D.J., and G. L. Uglem. 1984. Analysisofcontact-rehydration
other similar neurons, is positioned to control the syn- in terrestrial gastropods: absorption of l4C-inulin through the epi-
chrony of multiple behavioral responses normally ob- thelium ofthe foot../. Exp. Biol. Ill: 73-80.
served in response to environmental stress and changes Prior, D. J. 1989. Neuronal control ofosmoregulatory responses in
in the physiological state ofan organism. gastropods. Pp. 1-24 in Advances in Comparative Environmental
Physiology. R. Gilles, ed. 5. Springer-Verlag, New York.
Prior, D. J., and A. Gelperin. 1977. Autoactive molluscan neuron:
Acknowledgments reflex function and synaptic modulation during feeding in the ter-
restrial slug, Limaxmaximus J. Comp. Physiol. 114: 217-232.
parTt,hebywgorarnktsdferscormitbheedAirniztohinsapDaispeearsewaCosntsruoplpoRretseeda,rcihn PrioSLr.CimPDa.Bx,Ji.m,nacarxneidamsueWss..thHJe..rNWeeasuplrosonobsniio.vle.1n9e18s97s(.1o)f:tT8hh7ee-1fem0e5od.lilnugscmaontonreuprroopgeprtaimdeo,f
Commission (#82-0698) and The National Institutes of Prior, D.J.,and I. G. Welsford. 1989. Theroleofsmall cardioactive
Health (M.B.R.S.* 2 SO3 RR03401-03). peptide. SCPB. in the regulatory responses ofterrestrial slugs. Am.
Zool. 29: 1255-1263.
Prior, D.J., M.Hume, D. Varga,andS.D.Hess. 1983. Physiological
Literature Cited and behavioural aspects ofwater balance and respiratory function
in theterrestrial slug, Limaxmaximus. J. Exp. Biol. 104: 111-127.
Dickinson, P.S., D.J. Prior,andC. Avery. 1988. Thepneumostome Riddle,W.A. 1983. Physiologicalecologyoflandsnailsandslugs. Pp.
closure rhythm in slugs: a response to dehydration controlled by 431-461 in The Mollusca, W. D. Russell-Hunter, ed. Ecology. 6.
hemolymph osmolality and peptide hormones. Comp. Biochcm. AcademicPress, NewYork.
Physiol. 89A(4): 579-586. Schagene, K. A., I. G. Welsford, D. J. Prior, and P. A. Banta.
Furgal, S. M., and P. H. Brownell. 1987. Ganglionic circulation and 1989. Behavioraleffectsofinjection ofsmall cardioactivepeptide,
its effects on neurons controlling cardiovascular functions in the SCPB,on theslug Limaxmaximus. J Exp. Biol. 143: 553-557.
Aplysiacalilonuca J Exp. Zool. 244: 347-364. Teyke,T.,K.R.Weiss,and I. kupfermann. 1990. An identifiedneuron
Gelperin, A.,J.J.Chang,andS. Reingold. 1978. Feedingmotorpro- (CPR)evokesneuronal responsesreflectingfoodarousal inAplysia.
gram in Limaxmaximus:neuromuscularcorrelatesand control by Science247: 85-87.
chemosensory input.J. Neurobiol. 9: 285-300. Welsford, I.G.,and D.J. Prior. 1991. Modulationofheartactivityin
Grega, D. S., and D. J. Prior. 1985. The effects offeeding on heart the terrestrial slug Limax maximus by the feeding motorprogram,
activityintheterrestrialslug.Limaxmaximus:centralandperipheral small cardioactive peptides and stimulation of buccal neurone Bl.
control. J. Comp. Physio/. 156A: 539-545. ./. Exp. Biol. 155: 1-19.
Grega, D. S.,and D.J. Prior. 1986. Modification ofheart activity in Willows,A.O.D.,P.E.Lloyd,andB.P.Masinovsky. 1988. Multiple
response to dehydration in the terrestrial slug. Limax maximus. J transmitter neurons in Trilonia III. Modulation ofcentral pattern
Exp. Biol. 237: 185-190. generatorcontrollingpattern./ Neurobiol. 19(1): 69-86.
