AN ULTRASTRUCTURAL APPROACH TO FEEDING BEHAVIOUR IN PHILODINA ROSEOLA AND BRACHIONUS CALYCIFLORUS (ROTIFERS)

III. CILIA AND MUSCLES. CONCLUSIONS

P. CLEMENT, J. AMSELLEM, A.-M. CORNILLAC, A. LUCIANI’ & C. RICCI’

’ Laboratoire d’Histologie et Biologie Tissulaire (LA 244 CNRS) et CMEABG, Universite Claude Bernard (Lyon I), Villeurbanne, France; * Instituto di Zoologia, Universita degli studi, via Celosia, IO, Milano, Italy

Keywords: Rotifers, ultrastructure, feeding behaviour, cilia

Abstract

The orientation of the cilia pseudotrochus is controlled by mus- cles inserted on their rootlets. The peculiar structures of buccal and pharyngeal cilia are described: their beating leads the food towards the mastax. The circular and longitudinal muscles of the buccal funnel allow peristaltic movements and probably the rejection of food items not accepted by the mastax receptors.

Introduction

In Brachionus calyctjiorus and Philodina roseola, the acceptance or the rejection of food items seems to be made at least by the cilia of the pseudotrochus, the buccal funnel and the jaws of the mastax.

Optical microscope observations (Gilbert & Stark- weather, 1977 on B. calyciflorus) are insufficient to pre- cisely observe the structures involved in the feeding be- haviour. In the two preceding papers (Clement er al., I 980a and b) we described the buccal velum of Philodina roseola and Brachionus calyciflorus, and the oesophagial velum of B. calyczflorus: 6 is impossible to understand the structure and fish-trap function of these velums with an optical microscope.

Here we shortly describe the different effecters involved in the feeding behaviour: cilia and muscles. In conclusion, we try to make a synthetic scheme of the feeding be- haviour, involving both preceding papers, and also infor- mation not yet published about the nature and the loca- tion of sensory receptors.

The material and methods are the same as in the pre- ceding papers (see also Amsellem & Clement in this book). Our observations are more complete on Philodina roseola than on Brachionus calyciflorus.

Pseudotrochus

Cilia show the usual kinocil structure: 9 peripheral dou- blets, 2 central tubules, arms and intermediate structures; so they move actively and probably spontaneously.

They have, in their apical part, a cylindric dense struc- ture which has the diameter of a microtubule, beside two central tubules (Fig. I). The most interesting feature con- cerns the relation between the muscles and the ciliary rootlets. Each cilium is connected I) to the neighbour cilium by an horizontal network of striated ciliary roots and 2) to desmosomes by a striated ciliary root which goes down into the epithelial cell (Fig. I). The myofilaments of the longitudinal muscles are inserted on the other side of these desmosomes (Fig. I). Thus the muscles can control cilia orientation.

In Brachionus calyciflorus, the cilia are grouped: each group forms a cirrus, visible in optical microscopy.

Buccal funnel

The cilia here are also active kinocils. At their apical ex- tremity they have electrondense material (Fig. 2). A similar differentiation has been observed in Trichocerca rattus and Notommata copeus (Clement, r977a and b). Their striated rootlets go down to the perinuclear cyto- plasm (Fig. 2) but have no relation with muscles. The buccal funnel wall is formed by three layers:

I. the closely joined apical parts of the epithelial ciliary cells; they bear the cilia. Only this layer is schematised in Fig. 7.

2. a double muscle layer formed by an inner longitu- dinal layer and an outer circular one.

3. and finally the thickest layer which is made up of the ciliated epithelial cell bodies. These cell bodies are joined together by gap-junctions (Fig. 2).

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Hydrobiologia 73. I37-IdI(1980). oOI8-8158/80/0732-0137$OI.O0. 0 Dr. W. Junk b.v. Publishers. The Hague. Printed in the Netherlands.

Fig. I. 280~). Pseudotrochus cilia of Bruchionus calyciflorus. Axial section. The cilia(C) go through a thin cuticle (Cu). The striated ciliary rootlets (RC) insert on epitheliomuscular desmosomes (arrows). A longitudinal muscle (M) inserts on the other side of the same desmosomes. Pseudocoel (Ps). Encart. x ~ocmo. Transverse section of the extremity of a pseudotrochus cilium (Bruchionus calyczji’orus). A cylindric dense structure is juxtaposed to the two central tubules; Fig. Z. 13000. Transverse section of Philodina roseolu buccal tube. The circular (large arrows) and longitudinal (fine arrows) muscles are regularly and symmetrically disposed all around the tube. In the middle of the buccal lumen (BL),

138 the end of the buccal cilia contains dense-electron material. The rootlets (RC) of the buccal cilia are not attached to epitheliomuscular desmosomes: they originate inside the epithelial cell. The buccal velum (V) is in continuity with the

pharyngeal cells of the tube (p). All the epithelial cells bodies are linked by gap junctions (gi).

Fig. 3. x 6600. Transverse section of Philodin roseola mastax. The large curved arrows indicate the way taken by the food particles. The dorsal muscle (Mu) and the dorsal striated cilia(C) force them into contact with the uncus (un) and with the mastax sensory receptor (S); m: manubrium, M: mastax muscles; Fig. 4. x 6oooo. Axial section of three striated cilia of Philodina roseoZu (C in Fig. 3). Note their double cytomembrane and the localization of the dense transversal striated material, in the middle of the axonema; Fig. 5. x 60000. Axial section of two striated cilia of the mastax of Brachionus calyc1j7orus (C in Fig. 6). Note that the dense transversal striated material is situated between the axonema and the cytomembrane; Fig. 6. x 6600. Mastax of Bruchionus c4lyc1florus. Striated cilia(C) have rootlets (RC) which go down into the epithelial cell (EI) whose nucleus (NE) is partly visible. Two sensory neurites (sn) each have a tuft of I

sensory cilia situated among striated cilia. On the left, another epithelial cell (Ez) bears amyelin-like cuticle (Cu) which continues backwards to the cuticular oesophagus and forwards to the buccal velum.

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The regular disposition of the muscles (Fig. 2) indicates possible peristaltic movements in the buccal tube.

Pharynx

The pharynx is the part of the digestive tract located be- tween the buccal velum and the ciliary oesophagus (cf. the two preceding communications).

The pharyngeal cilia are found either just beneath the buccal velum or in a dorso-lateral position on the roof of the mastax (Fig. 7). They are short and have a larger di- ameter than those of the buccal funnel. They are active kinocilia (cf. Fig. IO in the communication I on the buccal velum). In Philodina roseola, they are surrounded by two cytomembranes.

The cilia located on the roof of the mastax show a specialization previously unknown anywhere else in the animal kingdom: they contain a periodically striated dense material (Fig. 4 and 5; Cornillac et al., 1979). This striated material is axial in Philodina roseola and periph- eral in Brachionus calyciflorus.

The most important muscles are located in the mastax: they insert on the trophi (Fig. 3 and 7) and are innervated by the ganglion of the mastax (Fig. 7). There is also a muscular layer around the mastax, inserted on the mastax and partly on the anterior cells of the rotatory apparatus. This muscular layer is present around the roof of the mastax (Fig. 3 and 7); its innervation comes from the brain (Fig. 7).

Localization of the sensory receptors involved in feeding behaviour

In Brachionus calyctji’orus, Clement (Ig77a and b) de- scribed chemoreceptors localized in the supple integu- ment between the lorica and the anterior cilia of the rotatory apparatus. Clement (1g77a and b) also described mechanoreceptor cilia which form tactile cirri in males and females of Brachionus calyciflorus.

Different receptors are also present in the anterior part of the body of Philodina roseola. In this species, no sen- sory receptor can be observed in the buccal tube. The only sensory structures between the pseudotrochus and the ciliary oesophagus are localized in the mastax (Fig. 7): two ciliary tufts among the periodically striated pharyn- geal cilia on the roof of the mastax, and a more complex receptor on the floor of the mastax, between the trophi

(Fig. 3 and 7). These sensory cilia seem to be chemo- receptors, and perhaps also mechanoreceptors (Clement et al., 1979).

In Brachionus calyciflorus, we find also two tufts of sensory cilia among the striated pharyngeal cilia (Fig. 6).

Conclusions on the phases and the mechanisms of the feeding behaviour in Philodina roseola and Brachionus calyciflorus (Fig. 7)

I. Food particles meet the anterior receptors of the animal by the beating of the cilia of the pseudotrochus.

2. The sensory structures are probably mechano- receptive as well as chemoreceptive. Their neurons are cerebral.

3. The brain innervates muscles which are inserted on the pseudotrochus ciliary rootlets: a first selection can be made because the cilia may allow or not allow the en- trance of food into the buccal funnel.

4. The food which goes through the buccal funnel then goes to the mastax owing to the action of buccal cilia, the buccal velum and the anterior pharyngeal cilia.

5. The mastax receptors (chemo- and perhaps also mechanoreceptors) sense the arriving food and release the trophi movements.

6. The food is pounded, filtered by special mastax cilia, then goes to the oesophagus. During the pounding and the filtration, mastax receptors can be informed by chemoreception if the food is acceptable or not and must thus be rejected or not. So, two types of behaviour are possible:

7. a) The food is considered suitable: the whole process runs on.

7. b) The food is considered unacceptable: the mastax receptors inform the mastax ganglion and the brain inner- vating the muscles of the mastax and those surrounding the buccal funnel: the mastax is projected forward to stop the food coming into the pharyngeal lumen, and the buccal funnel muscles reject this food towards the mouth owing to peristaltic contractions.

It seems that the food particles which first released the process in P. roseola are not rejected by this mechanism, because of the buccal velum position.

In rotifers, every muscle is innervated (Clement, 1g77a

and b); but we never observed an innervation of the ciliary epithelial cells.

So, in these conclusions, we suppose that ciliary beats are autonomous and not controlled by the nervous

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Fig. 7. Diagram of phases and mechanisms of the feeding be- haviour in Philodina roseola (and perhaps in Brachionus calyci- jlorus).

See the texrfor the descriptions of the dlfferentphases. There are three kinds of epithelial cilia:

- cilia of the pseudotrochus: in the pseudotrochus - buccal cilia with their dense apical part: in the upper part of the

buccal tube - very special and periodically striated cilia of the pharynx: on

the roof of the mastax. The longitudinal retractor muscles (MI) are inserted on the

rootlets of the cilia of the pseudotrochus. Longitudinal and circular muscles (Mz) surround the buccal

and pharyngeal cells in the buccal tube. All these muscles, and those located around the dorsal (M3) and ventral (M4) part of the mastax, and around the cuticular oesophagus (M6) are inner- vated by the brain. The muscles (MS) inserted on the trophi of the mastax are innervated by the mastax ganglion.

The different sensory receptors which seem involved in this behaviour are: I Anterior mechanoreceptors z Anterior chemoreceptors 3 Sensory receptors of the dorsal wall of the mastax 4 Sensory receptors of the ventral part of the mastax.

system, except in the pseudotrochus: at this level, the nervous system controls the cilia by way of the longitudi- nal rootlets inserted on the striated rootlets of the cilia.

These conclusions (Fig. 7) are a series of hypotheses based on ultrastructural observations; it is now necessary to test them by direct behavioural observations.

References

Amsellem, J. & Clement, P. r98oa. A simplified method for the preparation of rotifers for transmission and scanning electron microscopy. In this volume, pp. I 19-122. Rotifer Symp., Hydrobiologia.

Clement, P. r977a. Introduction a la photobiologie des rotiferes dont le cycle reproducteur est control& par le photoperiode. Approches ultrastructurale et exptrimentale. These no 7716. Universite Claude Bernard Lyon 1, France.

Clement, P. 1977b. Ultrastructural research on rotifers. Arch. Hydrobiol. Beih. 8: 270-297.

Clement, P., Cornillac, A. M. & Luciani, A. 1979. Cils chemo- recepteurs dans le mastax des Rotiferes. Biol. cell., 35, p. 2ga.

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Clement, P., Cornillac, AM., Amsellem, J., Luciani, A. 8r Ricci, C. I 980b. An ultrastructural approach to the feeding behaviour of Philodina roseola and Brachionus calyciflorus (Rotifers) II. The oesophagus. In: H. J. Dumont 8t J. Green (eds.) Proc. II Int. Rotifer Symp., Hydrobiologia.

Cornillac, AM., Luciani, A. & Clement, P. 1979. Des cils au con- tenu dense strit ptriodiquement, Biol. Cell., 35:

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