Inhibitory motor neurones supply body wall muscles in the locust abdomen
Neurobiologie, Universität Ulm, D-89069 Ulm, Germany
* Author for correspondence (harald.wolf{at}biologie.uni-ulm.de)
Accepted 17 October 2002
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Summary |
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In each abdominal ganglion half, there are two inhibitory motor neurones, CIa and CIb, which supply dorsal (CIa) and ventral (CIb) longitudinal muscles. Their cell bodies are located in the next anterior ganglion to where the axons leave the ventral nerve cord via nerve 1. Both inhibitors have contralateral somata in the posterior ventral soma cortex, looping primary neurites and bilateral dorsal arborisations. There are homonomous (segmentally homologous) motor neurones in the fused abdominal neuromeres, the thoracic ganglia, and at least the third subesophageal neuromere.
These body wall inhibitors are distinctly different from the limb muscle inhibitors, CI1-3, described previously. This is signified, for example, by the fact that both types of inhibitory motor neurones coexist in the prothoracic segment and innervate leg and body wall muscles, respectively.
Key words: locust, Locusta migratoria, common inhibitory motor neurone, abdominal muscle, segmental homology (homonomy), GABA
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Introduction |
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On the one hand, inhibitory motor neurones might well be reduced or absent
in body segments without appendages. Elimination of appendages and their
musculature in the course of evolution also abolished the need for the
neuronal control of these structures. Accordingly, the complement of neurones
which constitute the segmental ganglia in the abdomen is considerably reduced
when compared to the thoracic, limb-bearing segments. Even whole neuroblasts,
and thus the neuronal progeny generated by them, are missing from the
abdominal ganglion primordia in insects (e.g.
Shepherd and Bate, 1990).
Indeed, the neuroblast which gives rise to two of the three limb muscle
inhibitors in the thorax (Wolf and Lang,
1994
) is among those missing in the abdomen. Accordingly,
inhibitory motoneurones, where present in the abdomen, may not be homologues
of the inhibitors innervating the limb muscles.
On the other hand, the peculiarities of arthropod muscle that require
inhibitory control, namely sparse and polyneural innervation with (partly)
overlapping motor units (for reviews, see Rathmayer,
1990,
1998
), are the same in limb
and body wall muscles. Inhibitory motor neurones speed up the mechanical
response of a muscle to changes in excitatory motor neuron discharge through
the selective pre- and postsynaptic inhibition of `slow' motor neuron
activity, and through its effects on the muscle fibres
(Rathmayer and Erxleben, 1983
;
Rathmayer, 1998
). Common
inhibitors therefore facilitate rapid alternating limb movements, such as
executed during fast walking (Ballantyne
and Rathmayer, 1981
; Wolf,
1990
,
1992
). And although the
adjustment of muscle contraction properties according to changing behavioural
requirements is perhaps less demanding in body wall as opposed to walking leg
muscles, inhibitory innervation of body wall muscles would still appear
necessary where movements in distinctly different velocity domains are
executed, for instance, postural control and ventilation.
It was our goal, therefore, to ascertain whether or not inhibitory motor
neurones innervate the body wall muscles of the insect abdomen, and to
identify and describe possible inhibitory motor neurones. We combined extra-
and intracellular electrophysiology, labelling of motor neurones by peripheral
nerve backfills, and immunocytochemistry directed against the inhibitory
transmitter -aminobutyric acid (GABA) to identify inhibitory motor
neurones in the abdominal ganglia of the locust, Locusta migratoria
L. The general findings were verified in all segments between the last
subesophageal and the last unfused abdominal ganglia, although the fifth and
sixth abdominal ganglia were studied in particular detail. We discuss
segmental homology (homonomy) of inhibitory body wall motor neurones in the
locust ventral nerve cord, and homologies among the orthopteran insects.
We demonstrate that in the locust there are two (in the prothoracic
ganglion, three) body wall inhibitors per segment. Their morphologies are
consistent throughout the ventral nerve cord, with cell bodies located in the
anteriorly adjacent ganglion and contralateral to the nerve of axon exit,
posterior and ventral in the soma cortex, with looping primary neurites and
bilateral dorsal arborisations. Their targets are several ventral and dorsal
intersegmental muscles, most of which have segmental homologues (homonoms)
throughout abdomen and thorax. These inhibitory motor neurones of body wall
muscles are distinctly different from the limb muscle inhibitors examined
previously (Pearson and Fourtner,
1973; Hale and Burrows,
1985
; Watson et al.,
1985
; Wiens and Wolf,
1993
). In the thoracic ganglia, both types of inhibitory motor
neurones coexist and innervate the leg and body wall muscles,
respectively.
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Materials and methods |
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Locusts were opened by a dorsal midline incision, and gut, fatty tissue and dorsal and ventral diaphragms were removed. Usually head and thorax were discarded and the abdomen pinned open on a Petri dish coated with Sylgard (®, Dow Corning) (only in experiments where a normal respiratory rhythm was desirable were the more anterior body segments left attached). Where necessary for intracellular recording, the attachment sites of intersegmental muscles were fixed with minuten pins. The connectives were severed to isolate abdominal ganglia (AG) 4-7, and all nerves of AG 4, 5 and 7 were cut. This allowed ganglion 5 to be turned upside down, without strain to the remaining nerves, such that the ventral side was uppermost on a Sylgard-platform, with the nerve stumps immobilised with minuten pins. After digestion treatment with Protease (Sigma) for about 30s the ventral ganglion surface was accessible for intracellular recording from neuron cell bodies. The experimental arrangement is outlined in Fig. 1.
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The preparation was immersed in hypotonic locust saline, containing (in mmol l-1): NaCl 140, KCl 10, CaCl2.2H2O 2, NaH2PO4.H2O 4, Na2HPO4.2H2O 6, during preparation and experiments.
Electrophysiology
Extracellular en passant nerve recordings were made with single
hook electrodes, placed on the nerve of interest and insulated with
Vaseline.
Intracellular neuronal recordings were made from motor neuron somata in the
preparation described above. Electrodes made from thick-walled borosilicate
glass were used and had tip resistances of 80-160 M (tip solution, 3%
neurobiotin (Vector) in 1 mol l-1 KCl; shaft solution, 1 mol
l-1 KCl). The neurobiotin was injected with pulses of positive
current (2-5 nA, 1 Hz and duty cycle 50% for 30 min, or continuous current for
5-10 min). After fixation of the ganglion, neurobiotin was revealed using
Cy3-conjugated streptavidin (Sigma).
Intracellular muscle recordings were performed with flexible electrodes
according to Pearson and Iles
(1971). Thin-walled
borosilicate glass electrodes with tip resistances of 20-40 M
were
filled with 3 mol l-1 potassium acetate. Shafts were shortened to
about 1 cm length and mounted on Ag/AgCl-wires (100 µm diameter) with
silicone elastomer (Kwik-Cast, WPI). Recordings were made near the middle of
the muscle fibre.
In experiments where just the presence of inhibitory muscle innervation was to be examined, the preparation was usually left at 4-8°C for 1-2 h before experiments. During this time, the resting membrane potential of the muscle fibres stabilised at between -20 and -40 mV, depending on muscle fibre type. This was probably due to reduced oxygen supply after removal of the main tracheae. In this situation, inhibitory postsynaptic potentials (IPSPs) were clearly discernible. The activity of excitatory motor neurones was reduced and often absent, minimising muscle movement, while the inhibitory motor neurones showed increased spontaneous activity.
Immunocytochemistry
The immunocytochemical procedures were adapted after Stevenson et al.
(1992,
1994
) and are outlined only
briefly, except where they were different from standard procedures. The
nervous system was dissected from the animal and immediately fixed in
glutaraldehyde/picric acid/acetic acid solution (GPA;
Boer et al., 1979
) for 2 h at
room temperature. Following dehydration and rehydration in an ethanol series
(50, 70, 90, 96 and 100%), the tissue was washed in 0.1 mol l-1
phosphate-buffered saline (PBS) containing 0.1 mol l-1 phosphate
buffer, pH 7.4, 0.1 mol l-1 NaCl; washing steps were also
interspersed between the following incubations. To saturate double bonds
(Stevenson et al., 1994
) the
ganglia were incubated in 0.5% NaBH4 in PBS for 30 min. Then they
were digested in a collagenase/dispase, hyaluronidase mixture (1 mg
ml-1, Roche and Sigma, respectively) for 30 min to 2 h, depending
on the preparation, at 37°C to improve access for antibodies. Following 1
h incubation in 10% normal goat serum (NGS) in PBS with 0.5% Triton-X-100
(TrX, Sigma), the nervous system was incubated in 1:10 000 diluted GABA
antiserum (polyclonal antibody raised in rabbit, SFRI, France) in PBS with
0.1% TrX and 1% NGS for 48 h at 37°C. Incubation with the secondary
antibody (Cy2-conjugated goat-anti-rabbit IgG, or biotinylated
goat-anti-rabbit IgG, both from Dianova) was in PBS with 0.1% TrX for 12 h at
37°C, and at 1:100 dilution. After washing for 24 h at 37°C, the
nervous system was dehydrated, cleared in methylsalicylate and embedded in DPX
(Fluka) for subsequent microscopic examination. The specificity of the primary
antibody was tested at regular intervals according to standard protocols
(mainly omitting the primary antibody, testing different fixatives, etc.).
Backfills and histology
The motor neuron supply of selected muscles was labelled by backfilling
with 3% neurobiotin in distilled water. The motor nerve was immersed in the
filling solution for about 12 h at 4-8°C. The marker substance was
contained in a small vaseline well located close to the muscles in question.
Individual motor neurones were filled with neurobiotin by intracellular
impalement and current injection (see above). Following selective labelling by
nerve backfill or intracellular injection, neurobiotin was coupled to
Cy3-conjugated streptavidin. As for the immunocytochemical preparations, the
backfills were first examined and photographed under a confocal
epifluorescence microscope (DMRE/TCS SP, Leica). Further histological
processing was conventional, and ganglia were sectioned at 8-10 µm for
reconstructions (e.g. Fig.
7).
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Results |
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Electrophysiology
A combination of extra- and intracellular electrophysiology was employed
for the initial examination of a possible inhibitory innervation of body wall
muscles in the locust abdomen. Fig.
1 illustrates the experimental situation. Prominent inhibitory
postsynaptic potentials (IPSPs) were recorded in several body wall muscles,
namely, dorsal, pleural and ventral intersegmental muscles. In abdominal
segment 6, these were muscles 212a,b,c,d, 213a,b (dorsal intersegmental
muscles), 214 (pleural intersegmental muscle), 217 and 218 (ventral
intersegmental muscles), nomenclature according to Snodgrass
(1935). In the other abdominal
segments with unfused ganglia these were the segmentally homologous
(homonomous) muscles (see Table
1). The amplitudes of IPSPs markedly decreased with more negative
membrane potentials imposed by current injection, attesting to the inhibitory
innervation of these muscles (reversal potentials were not determined,
however). Simultaneous extracellular nerve recordings demonstrated that the
axons of the inhibitory motor neurones left the ventral nerve cord through
nerve 1 of the segmental abdominal ganglion. Note that muscles 219, 220, 221,
222 and 224, and their segmental homologues (homonoms) in other segments, did
not receive inhibitory innervation according to all our experiments (see also
below).
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Intracellular recordings from motor neuron somata were guided by the results of combined motor neuron backfills of nerve 1 and immunocytochemistry directed against the transmitter of insect inhibitory motor neurones, GABA (see below). Fig. 2 shows an intracellular recording from the cell body of an inhibitory motor neuron (CIa), a simultaneous extracellular en passant recording from nerve 1 of the segmental ganglion (N1AG6), and an intracellular recording from a dorsal intersegmental muscle (M212a). Five sample traces are superimposed, illustrating the 1:1 correspondence of action potential recordings in the motor neuron soma and nerve 1, respectively, and IPSPs in the muscle fibre. This recording provides proof of inhibitory innervation of muscle 212a by motor neuron CIa. Inhibitory innervation of every muscle shown in Fig. 1 was tested and verified in this way. The time delay between centrally recorded motor neuron spikes in CIa and IPSPs in muscle 212 was approximately 20 ms, equivalent to a conduction velocity of approx. 0.7 m s-1.
|
In other body segments, inhibitory muscle innervation was often examined in a more simple way. IPSPs were recorded in one muscle (usually a readily accessible dorsal intersegmental muscle) and the supply of other muscles by the same inhibitory motor neuron was sought by examining recordings from nerve 1 and from the nerve branch supplying a particular muscle, and sometimes also by a second intracellular muscle recording. A 1:1 correspondence of spikes and IPSPs was taken as strong indication of inhibitory innervation by the same neuron. Immunocytochemical data (see below) supported these electrophysiological results. In summary, the homonoms of all the muscles (shown in Fig. 1 for the sixth abdominal segment) were found to be supplied by their respective inhibitory motor neurones in abdominal segments 1-7. In the thorax, too, a comparable innervation pattern was observed. The data are summarised in Table 1 and discussed in detail below.
Identification of abdominal inhibitory motor neurones CIa and
CIb
The combined recording from nerve supply, muscle fibre and inhibitory motor
neuron described above (Fig. 2)
was used to identify neurones subsequently injected with neurobiotin to reveal
their morphologies (Fig. 3B;
CIa, N=4; CIb, N=7). Fig.
3A demonstrates the 1:1 correspondence of spikes in the cell body
recordings (top traces) and IPSPs in the impaled muscle fibres (bottom traces)
(nerve recordings, middle traces). Fig.
3C illustrates that, prior to labelling, identification of the
inhibitory motor neuron cell body was substantiated by depolarising current
injection, which allowed both a clearer assessment of the 1:1 correspondences
noted above, and direct control of IPSP generation in the muscle fibre. Note,
for instance, the summation of IPSPs in muscle 212a (third trace from top) due
to the high spike frequency in CIa (most clearly seen in the recording from
nerve 1, second trace).
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In each abdominal segment, two inhibitory motor neurones that supplied the
body wall muscles could always be identified. Since both innervated two or
more muscles (Fig. 1), they are
Common Inhibitory motor neurones. Hence, we termed them CIa
(Fig. 3Ai,Bi) and CIb
(Fig. 3Aii,Bii), to provide a
clear distinction (see below) from the thoracic common inhibitors,
CI1-3, described previously (e.g.
Hale and Burrows, 1985). The
cell bodies of both motor neurones resided in the ganglion anterior to the
position where their axons exited the central nervous system in nerve 1
(Fig. 1). They were located in
the posterior ventral cortex of the ganglion, usually just contralateral of
the midline, although the exact soma positions were quite variable.
Occasionally, CIa soma was on, or slightly ipsilateral to, the midline. The
soma of CIa was between 25 and 30 µm in diameter, and thus distinctly
larger than the soma of CIb, which measured about 20 µm across. The path of
the primary neurite made a very characteristic loop, running dorsolaterally
from the soma towards dorsal commissure V, (DCV;
Watson and Pflüger,
1987
), CIa with a sharp turn, and CIb far into the contralateral
hemiganglion with a rounded loop (Figs
3Bi,ii,
7). The dendritic arborisations
covered nearly the complete dorsal neuropil area of the ganglion and were
almost bilaterally symmetrical. Only CIa had a few minor dendrites extending
ventrally. The two halves of the bilateral arborisations were connected at the
ganglion midline, close to where the primary neurite joined them. Usually
three or four prominent secondary neurites were discernible per hemiganglion
(Fig. 3Bi). CIa always
exhibited more profuse and dense arborisations than CIb. The main neurites of
CIa and CIb entered dorsal commissure V, and their axons left the segmental
ganglion through the posterior connective to exit the ventral nerve cord
through nerve 1 of the next more posterior ganglion (arriving there from the
dorsal intermediate tract, DIT) (Fig.
7). In keeping with its larger cell body, CIa also had the larger
axon, even in the peripheral nerves.
Activity patterns of the abdominal inhibitors
Once CIa and CIb had been identified, their target areas (see above) and
activity patterns could be characterised in more detail.
Fig. 4 illustrates the
discharge patterns of both inhibitors, by means of recordings from muscle 212,
supplied by CIa, and muscle 217, supplied by CIb (a recording from nerve 1
monitored overall neuronal activity). The IPSPs generated in the muscle fibres
by the inhibitory motor neurones are clearly visible, reflecting the
inhibitors' spike activities (compare Fig.
2). The (larger and briefer) depolarisations produced by
excitatory input are also visible; however, they were not studied here (see
e.g. Thompson et al., 1999).
It is evident that both CIs discharged their highest spike frequencies just at
the beginning of the inspiration phase, perhaps actively terminating muscle
contractions needed for expiration. Spike activity in CIb was restricted to
the beginning of the inspiration phase, while CIa also produced action
potentials during the later part of the inspiration and the beginning of the
expiration phases, albeit at much lower frequencies. Spike activity in CIa
was, on average, higher and also reached higher peak frequencies than the
discharges in CIb. CIa activity ceased reliably only around the middle of
inspiration.
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Immunocytochemistry
Anti-GABA immunocytochemistry was employed to mark the cell bodies of
putative inhibitory neurones in the ganglia of the ventral nerve cord, and
putative inhibitory axons in the peripheral nerves (see Materials and
methods). Combining anti-GABA immunocytochemistry with backfills of nerve 1
provided another method to identify inhibitory motor neurones CIa and CIb
(Fig. 5). The backfill
technique labelled motor neurones with axons in nerve 1
(Fig. 5Ai,Bi,Ci), while
immunocytochemistry labelled the cell bodies of GABA-immunoreactive, and thus
putative inhibitory, neurones (Fig.
5Aii,Bii,Cii), including motor neurones. Combined staining thus
identified inhibitory motor neurones with axons in nerve 1
(Fig. 5Aiii,Biii,Ciii). There
were always (N=58) two inhibitory motor neuron somata labelled in the
abdominal ganglion anterior to the nerve 1 backfill. According to the
electrophysiological data described above these were unequivocally identified
as the cell bodies of CIa and CIb. Soma size and location were in agreement
with those given in the anatomical description above.
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In the nerves 1 of all unfused abdominal ganglia there were two
immunoreactive axons, which obviously entered the segmental ganglion from the
ipsilateral anterior connective and again left through the nerve root before
making contact with any structures in the ganglion itself (arrows in
Fig. 5Aii, Aiii). In a few
cases (N=11), it was confirmed through intracellularly recorded and
dye-injected inhibitory motor neurones that in abdominal ganglion 6, the
thicker of the two axons was that of CIa, while the thinner axon belonged to
CIb. In segments where abdominal neuromeres are fused to each other (A8-10,
A11 not yet clear) or to the metathoracic ganglion (A1-3), and also in the
thoracic ganglia (T1-3) and the third subesophageal neuromere (S3),
corresponding immunoreactive axons were observed in the nerve roots, which
contain the axons of neurones homonomous to those running through abdominal
nerve roots 1 (Steffens and Kutsch,
1995) (summarised in Fig.
8).
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When the two immunoreactive axons in abdominal nerve 1 were followed into
the periphery, they were observed to innervate the muscles noted in
Fig. 1 for abdominal segment 6.
This is illustrated for muscle 218 and the axon of CIb in
Fig. 6. Even synaptic boutons
were visible in many immunocytochemistry preparations, usually on the more
central fibres of a given muscle, and in agreement with the
electrophysiological muscle fibre recordings reported above, where inhibitory
innervation was often absent in the superficial fibre layers of a muscle.
These observations are in agreement with previous accounts of arthropod muscle
structure (e.g. Müller et al.,
1992), stating that CI innervation parallels innervation by `slow'
motor axons, and that the muscle fibres supplied by `slow' axons usually
occupy the central portion of a muscle
(Rathmayer and Erxleben,
1983
). Above all, these observations confirmed the results of the
electrophysiological experiments described above, namely, inhibitory
innervation of muscles 212, 213, 214, 217 and 218 in abdominal segment 6
(Fig. 1), and of the homonomous
muscles in the other abdominal segments. Inhibitory innervation was evidently
absent in muscles 219, 220, 221, 222 and 224, and their homonoms.
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Immunocytochemistry also indicated that a number of body wall muscles in the thorax receive inhibitory innervation from motor neurones homonomous to CIa and CIb (summarised in Table 1), although CIa and CIb were rigorously identified by intracellular electrophysiology only in the sixth abdominal segment.
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Discussion |
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At a first glance, the presence of inhibitory motor neurones in the
abdominal segments of the locust may appear surprising. Considering the
absence of appendages (except genitalia and cerci in the most posterior
segments), and the fact that almost all insect inhibitory motor neurones
described so far subserve the control of appendicular musculature, one might
have expected the absence of inhibitory muscle innervation in the abdomen (see
Introduction). Walking legs (Hale and
Burrows, 1985), antennae
(Honegger et al., 1990
), and
even crustacean mandibles (Ferrero and
Wales, 1976
) are all supplied by common inhibitory motor neurones.
Whether there is a similar inhibitory supply of body wall muscles was as yet
unknown, except for a few cursory reports for locust
(Yang and Burrows, 1983
;
Bräunig, 1993
;
Bräunig et al., 2002
;
Schmäh et al., 2001
;
Schmäh and Wolf, 2002
),
cricket and bushcricket (Consoulas and
Theophilidis, 1992
; Consoulas
et al., 1993
). The body wall inhibitors, CIa and CIb, are not,
however, homologues of the common inhibitors which supply the leg muscles in
the thorax, CI1-3. This is most clearly demonstrated by the
coexistence of both types in the prothoracic segment
(Bräunig et al., 2002
;
Schmäh, 2002
;
Schmäh and Wolf, 2002
),
where they supply body wall and leg muscles, respectively. The two classes of
inhibitors probably have different developmental origins, since the
neuroblast, which gives rise to two of the leg inhibitors, neuroblast 5.5
(Wolf and Lang, 1994
), is
absent in the abdominal ganglia. They nevertheless share several common
features, namely, the innervation by each neurone of several muscles, the
contralateral, posterior and ventral position of the motor neuron somata, and
some characteristics of the trajectories of the primary neurites within the
neuropil (Fig. 7).
It is not clear at present whether or not these similarities extend to the
inhibitors' function. Innervation of several muscles by a single motor neuron
would indicate a global function in setting muscle performance, as
demonstrated for the limb inhibitors (see
Wiens, 1989). The data
available on the activity patterns of body wall inhibitors
(Fig. 4) may support this
interpretation. CIa and CIb are most active just at the beginning of the
inspiration phase. This is also observed in the bushcricket
(Consoulas and Theophilidis,
1992
). Although the intersegmental muscles supplied by CIa and CIb
are not involved in the main (dorso-ventral) respiratory movements of the
abdomen, the inhibitors might still contribute to the termination of muscle
contractions supporting expiration, or to the fast execution of the initial
inspiration movements. As yet unknown are the activity patterns of CIa and CIb
during other abdominal motor tasks, namely, postural control at rest, walking,
flight and flight steering (Dugard,
1967
; Baader,
1988
), and during copulation and oviposition.
Homonomy (segmental homology)
Apparently, there are segmental homologues (homonoms) of the abdominal
inhibitory body wall motor neurones in other segments of the locust body
(summaries in Fig. 8,
Table 2), as indicated by
anti-GABA immunocytochemistry, and selected nerve (partly P. Bräunig,
personal communication) and muscle (partly T. Müller, personal
communication) recordings. When these data are combined with morphological
descriptions in previous studies
(Bräunig, 1993;
Bräunig et al., 2002
;
Kutsch and Heckmann, 1995
;
Steffens and Kutsch, 1995
;
Schmäh et al., 2001
;
Schmäh, 2002
;
Schmäh and Wolf, 2002
),
they provide an unequivocal identification of inhibitory motor neurones in the
following sections of the ventral nerve cord: (i) the third subesophageal
neuromere, (ii) the pro- and mesothoracic ganglia, (iii) the three abdominal
neuromeres fused with the metathoracic ganglion, and (iv) all unfused
abdominal ganglia. The presence of two homonomous inhibitory motor neurones in
the metathoracic ganglion and in the most anterior neuromeres of the terminal
ganglion are predicted only on the basis of the present immunocytochemical
data.
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In their backfill study of ventral body wall muscles, Steffens and Kutsch
(1995) provided detailed
anatomical descriptions of `common contralateral A-type cells' in all ganglia
and neuromeres between the third subesophageal neuromere and seventh abdominal
ganglion, except the metathoracic neuromere. These `common contralateral
A-type cells' exhibit all the characteristics of CIb, including common
innervation of ventral intersegmental muscles, contralateral and posterior
soma location, and looping primary neurite. Since the backfills of Steffens
and Kutsch (1995
), and our own
backfills of nerves 1 (Fig. 5),
marked the full complement of motor neurones that supply the respective
ventral body wall muscles, comparison of these data with the
immunocytochemical labelling identifies homonoms of CIb in the segments named
above. A similar line of argument holds for the `loop cell' described by
Kutsch and Heckmann (1995
),
which innervates dorsal body wall muscles. This cell has been examined in the
third subesophageal neuromere, the prothoracic ganglion, the mesothoracic
ganglion, and abdominal ganglia 2, 4 and 6. The `loop cell' is homonomous to
CIa by the criteria just noted.
In the case of the terminal ganglion, there are no previous studies that would allow homonomisation of equivalents of the `loop cells' or `contralateral A-type cells' with CIa or CIb. The present immunocytochemical data are strongly suggestive, however. There is the expected pattern of immunoreactive neurones in abdominal neuromeres 8 and 9, i.e. two axons each leave through the third nerve root of the terminal ganglion (tergal nerve of neuromere 9) and through the epiproct nerve (corresponding to nerve 1 of neuromere 10), and two immunoreactive motor neuron somata per hemineuromere exist in the appropriate regions of the soma cortex (indicated in Fig. 8). No inhibitory motor neurones appear to exist in neuromeres 10 and 11, which carry the genitalia, and no immunoreactive axons were observed in the corresponding nerves. Despite this consistent general pattern, there was sometimes variability, even asymmetry, in the courses of the inhibitory axons and even in the number of axons per nerve.
These findings are in good agreement with the immunocytochemical study of
Watson and Pflüger
(1987), except that these
authors occasionally observed thin immunoreactive structures in the cercal
nerve and even in nerves 2 of the eigth neuromere and the free abdominal
ganglia. Considering that two species of locust were examined, different
antisera were used in the two studies, and that GABA immunocytochemistry is
prone to false positive as well as false negative results, this difference is
not too surprising, and the issue awaits further scrutiny.
There are two immunoreactive axons in nerve 6 of the metathoracic ganglion
and two immunoreactive somata with the appropriate morphological
characteristics in the metathoracic neuromere, indicating a normal complement
of two body wall inhibitors in the metathoracic neuromere. In the prothoracic
ganglion, a third inhibitory motor neuron of body wall muscles is present, in
addition to the CIa and CIb homonoms, according to Bräunig
(1993), Bräunig et al.
(2002
), Schmäh and Wolf
(2002
), and the present
immunocytochemical data. The morphology of this third inhibitor is strongly
reminiscent of that of the two other body wall CIs, suggesting a close
relationship.
Species comparison and evolutionary considerations
The presence of three body wall inhibitors in the locust prothoracic
segment raises the question of whether or not there might be three inhibitory
body wall motor neurones in other insects, and what the original, evolutionary
plesiomorphic complement of body wall inhibitors might be
(Schmäh, 2002). The
situation in the abdominal ganglia of phasmids is similar to that in the
locust, and thus allows no further conclusions. In the abdominal ganglia of
dragonflies, by contrast, there appear to be three inhibitory motor neurones,
all with their major features reminiscent of CIa and CIb in the locust
(Schmäh, 2002
). Data from
crickets and bushcrickets point in a similar direction
(Consoulas and Theophilidis,
1992
; Consoulas et al.,
1993
; Böser,
1999
). Böser
(1999
) reports three inhibitory
motor axons, and associated cell bodies, which supply body wall muscles in the
pro-, meso- and metathoracic ganglia of the cricket Acheta
domesticus. As in the locust prothorax, these body wall inhibitors
coexist with the leg inhibitors, CI1-3. Although preliminary, these
observations, taken together, suggest that the original, plesiomorphic
situation in pterygote insects is indeed a complement of three body wall
inhibitors, a condition that is reflected in the locust prothoracic
ganglion.
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Acknowledgments |
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References |
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