Hydrobiologia 387/388: 135–140, 1998.E. Wurdak, R. Wallace & H. Segers (eds), Rotifera VIII: A Comparative Approach.© 1998 Kluwer Academic Publishers. Printed in the Netherlands.

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Catecholaminergic neurons in the brain of rotifers

Elena A. KotikovaZoological Institute of the Russian Academy of Sciences, 199034, St. Petersburg, Russia

Key words:nervous system, morphology, cathecholamines, GAIF method, Rotifera

Abstract

In 10 rotifer species from the subclasses Archeorotatoria (order Bdelloidea) and Eurotatoria (superorders Gnesio-trocha and Pseudotrocha) three patterns of catecholaminergic neurons are detected, namely: x-shaped, arch-shapedand ring-shaped. These brain complexes are developed independently and in a parallel fashion in different rotifergroups. The number of the brain catecholaminergic neurons varies from 6 to 11, constituting about 3–7% of thetotal number of the brain cells. The brain neuron pattern demonstrates a distinct bilateral symmetry.

Introduction

A general phylogenetic system of rotifers based onmorphological characters has not yet been elaborated(Kutikova, 1985), although this group should be re-garded as a separate phylum (Malakhov, 1994). Thesystem of phylogenetic relationships between the or-ders of rotifers based on mastax structure (Markevich,1990), is considered here to be the most realistic. The‘classical’ nomenclature is given in parentheses.

Electron microscopic investigation of the rotifernervous system revealed the presence of dense-coredsynaptic vesicles, which are supposed to containcatecholamines (CA) (Villeneuve & Clement, 1971;Clement, 1977; Wurdak et al., 1983). Later, histo-chemical studies have localized CA in whole-mountsof rotifers (Nogrady & Alai, 1983); Keshmirian &Nogrady, 1987, 1988).

The mapping of CA distribution in the nervous sys-tems of rotifers (Kotikova, 1994, 1995, 1997) shouldbe regarded as a next step in these studies.

The mapping of biologically active substanceswhich are characteristic of the simple nervous systemsof invertebrates, opens new opportunities to study in-vertebrate nervous system structure and origin in moredetail.

Materials and methods

10 species of Rotifera were studied:

subclass Archeorotatoria (order Bdelloidea)Order Bdelloidea

Philodinasp.subclass Eurotatoria (superorders Gneisiotrochaand Pseudotrocha P. Beauchamp, 1965)

order TransversiramidaEuchlanis dilatataEhrenbergManfredium eudactylotum(Gosse)Brachionus quadridentatusHermannPlatyias quadricornis(Ehrenberg)Lecane (Monostyla) arcuata(Bryce)

order SaltiramidaAsplanchna herrickiGuerneAsplanchna priodontaGosse

order SaeptiramidaNotommatasp.

order AntrorsiramidaDicranophorus forcipatus(Muller)

CAs were revealed by the glyoxylic acid in-duced fluorescence (GAIF) method slightly modifiedby Kabotyanskyj (1985) with an increased (up to5%) concentration of GA in the incubation mediumprepared with 0.1 M Na-phosphate buffer pH 7.2.

The controls were carried out in two ways: exclu-sion of the GA from the incubation medium and addi-tion of water to the dried preparations. Both reactionsresulted in disappearance of the fluorescence. Thegreen fluorescence of CA-containing elements was ob-served in a fluorescent microscope LUMAM-R3; thephotomicrographs were taken on RF film.

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Figure 1. Schematic drawing of the catecholaminergic nervous system revealed by the GAIF method in the rotiferAsplanchna herricki. A:View from the ventral side. B: View from the lateral side. BG – brain ganglion; E – eye; MN – mastax neuron; N – neuron; RGN – rudimentarypedal gland neuron; RN – urinary bladder neuron; VLC – ventro-lateral nerve cord; VT – varicose thickening.

Results

Asplanchna herrickiGuerne (Figures 1A–B; 2D)The brain ganglion looks like a sharply curved archon the dorsal side between the eyes and the pharynx.A pair of central neurons is situated on both sides ofthe cerebral eye spot. A second pair of larger neuronslies under the lateral eye spots. The processes of thethird pair of brain neurons bend to the ventral sideand give rise to the longitudinal ventro-lateral nervecords. These cords run along the sides of the body andpresent varicose thickenings along their length. A pairof bipolar neurons is located at the level of the distalpart of the stomach. The ventro-lateral cords join eachother near the urinary bladder. A pair of unipolar neur-ons which send their axons into the longitudinal cordis situated at the sides of the urogenital duct. A pairof small unipolar neurons with axons directed to thelateral parts of the brain arch is located just under thelateral eye spots.

A pair of unipolar neurons lies near the mastax.Their processes first run posteriorly along the mas-tax at the ventral side of the body and then contour

the mastax and end behind it by globular sensoryterminals.

Above the urogenital opening near a pair of rudi-mentary pedal glands lies a pair of small unipolarneurons with their processes oriented along the ducts.All the neurons are oval-shaped or elongate; their sizevaries from 2 to 6µm. The similar general patternof the CA-ergic nervous system is characteristic forA. priodonta(Figure 3D) and for other species de-scribed below. However, some noticeable variations inthe brain neuron pattern are observed (Figure 2).

Philodinasp. (Figures 2A, 3A)The X-shaped brain complex is situated above thepharyngeal tube behind the cerebral eye spots andincludes seven longitudinally oriented neurons. Theneurons lie at three levels. At the upper level a pairof widely separated large neurons is observed. Themiddle level includes a pair of bipolar perikarya withapposed processes. At the lower level three oval-shaped neurons are located. Peripheral processes ofthe first pair of brain neurons are directed anteriorlyand end at the base of corona by spherical sensory

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Figure 2. Scheme of the pattern of CA-ergic brain neurons and neurons of the corona in the rotifers: A –Philodina sp., B –Manfrediumeudactylotum, C – Platyias quadricornis, D – Asplanchna herricki, B – Euchlanis dilatata, F – Notommatasp., G –Lecane arcuata, H –Brachionus quadridentatus, I – Dicranophorus forcipatus. Dorsal neurons are shown in black; ventral neurons are shown in outline. Neuronsof the corona, of the anterior dorsal tentacle and the proboscis are indicated by small arrows.

terminals. Five neurons connected by their processesforming a ring-shaped complex lie around the dorsalproboscis. Three small neurons of this complex liedorsally and two larger ones are ventrally located. Thesize of the neurons is 2–7µm.

Manfredium eudactylotum(Gosse) (Figures 2B, 3B)

The brain complex of CA-ergic neurons looks like aslightly curved X-shaped structure, situated just be-hind the eye spot. It is formed by eight neurons. Theneurons are longitudinally extended on four levels.The neurons of the two upper levels are larger (6µm),those of lower levels do not exceed 3µm.

Platyjas quadricornis(Ehrenberg) (Figures 2C, 3C)

The arch-shaped brain ganglion is gently curved. Itincludes seven bipolar neurons (5–7µm) forming atransverse row (Kotikova, 1995).

Euchlanis dilatataEhrenberg (Figures 2E, 3E)

The brain arch is characterized by a conical curve andconsists of six bipolar neurons (4–6µm).

Notommata sp.(Figures 2F, 3F)

CA-ergic brain neurons are arranged in a semicircu-lar pattern. Four neurons form a nearly straight dorsalsector, two neurons (6–8µm) are shifted to the ventralside and lie at the posterior end of the ganglion at thelevel of the eye spot. Axons of the two unipolar neur-

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Figure 3. CA-ergic neurons in the brain of rotifers: A –Philodina sp., B –Manfredium eudactylotum, C – Platyias quadricornis, D –Asplanchna priodonta, E – Euchlanis dilatata, F – Notommatasp., G –Lecane arcuata, H – Brachionus quadridentatus. GAIF method.Scale bar = 10µm. Brain neurons are indicated by small arrows.

ons (4µm) lying at the level of the corona approachthe lateral parts of the dorsal arch.

Lecane (Monostyla) arcuata(Bryce) (Figures 2G,3G)Brachionus quadridentatusHermann (Figures2H, 3H)These species are characterized by a ring-shaped brainganglion. L. arcuata has a more compact ganglionwhich is interrupted ventrally, whileB. quadridentatushas a ganglion extended in transverse direction. It in-cludes six neurons (4–8µm), four of which are locatedon the dorsal side and two others lie on the ventralside. InL. arcuatatwo nerves of the corona with two

pairs of neurons (4–7µm) originate from the dorsalpart of the brain. A pair of small unipolar neuronssends short axons to the dorsal part of the brain. InB.quadridentatus, a pair of small bipolar neurons runsfrom the medial part of the dorsal brain semicircle tothe anterior dorsal tentacle.

Dicranophorus forcipatus(Muller) (Figure 2I)Eleven CA-ergic neurons form a ring-shaped brainganglion in the posterior part of the head region. Atransverse dorsal part of the brain includes three mainneurons and four additional ones while the curvedventral part has four neurons. Four unipolar neurons

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lie at the level of the frontal eye spots, a pair of dorsalneurons among them sends axons into the brain, whilethe ventral neurons have no processes. The size ofneurons varies from 2 to 7µm.

Discussion

All the rotifers studied, being systematically distantfrom each other, nevertheless show the same patternof the CA-ergic nervous system. Main differences areobserved in the location of the brain neurons, theirnumber varying from 6 to 8 (without considering theadditional perikarya, which have no influence on theganglion shape). Three geometric patterns of CA-ergic brain neurons can be distinguished: X-shaped,arch-shaped and ring-shaped. Brain neurons can belongitudinally extended at three or four levels form-ing X-shaped figures of different degrees of curvature(Philodinasp.,M. eudactylotum). Among the specieswith the arch-shaped brain, a gentle curve is recordedin P. quadricornis, butA. herricki, A. priodontaandE.dilatata are characterized by a steeper curve. Trans-ition from the arch-shaped to the ring-shaped brainis realized through two stages. At the first stage aninsignificant degree of curvature is shown inNotom-mata sp. and a further approachment of the lateralbrain parts is observed inL. arcuata. The second stageleads to the appearance of the closed ring-shaped braincomplex ofB. quadridentatusandD. forcipatus. Suchstructural changes seem to take place independentlyin different groups and result in the appearance of asimilar geometric pattern of CA-ergic brain neuronsin representatives of different subclasses (Philodinasp. andM. eudactylotum). On the other hand, sim-ilar patterns can be observed in the representatives ofthe same order Transversiramida (E. dilatata and P.quadricornis) as well as among the members of thethree orders Saeptiramida, Transversiramida and An-trorsiramida (Notommatasp., B. quadridentatus, D.forcipatus) which are not closely related to each other.

Among the Archeorotatoria (Bdelloidea) onlyone pattern of brain CA-ergic neurons is described(Kotikova, 1994, 1995), while among the Eurotat-oria three patterns of it have been revealed (Kotikova,1995, 1997).

Numerous similarities in forms of movement,feeding, structure of the corona, general morphologywere described for rotifers (Kutikova, 1983). The ex-cretory system demonstrates distinct examples of suchparallelisms (Remane, 1929–32).

The pattern of CA-ergic elements ofAsplanchnaandBrachionusfalls into the classic scheme of theirnervous system (Nachtwey, 1925; Stossberg, 1932).The CA-ergic fibres are not detected in the dorsal lon-gitudinal cords ofAsplanchna priodonta(Nachtwey,1925) indicating that these cords have only a motorfunction. Similar cords can be expected to be found inother species of rotifers.

Brain neurons of rotifers constitute 20% of the totalnumber of cells (Martini, 1912). The brain ofAs-planchna brightwellinumbers 200 cells, demonstrat-ing a perfect bilateral symmetry (Ware & Lopresti,1975). The distinct bilateral symmetry is shown alsoin the arrangement of CA-ergic neurons constitutingin different species only 3–7% of the total number ofcells in the brain.

The pattern of CA-ergic nervous system of roti-fers, once it evolved, was maintained during the wholeperiod of evolution of the group. A high degree ofconcentration of CA-ergic elements can be regardedas an indicator of the high level of development of thesimple nervous system of rotifers.

Acknowledgements

This work was supported by the Russian Basic Re-search Foundation (grant N96-04-48384). I want toexpress my gratitude to L. Kutikova and G. Markevichfor their help in the identification of rotifers and also toA. Kirejtshuk and to O. Raikova for their help in trans-lation of the text. My sincere thanks to the OrganizingCommittee of the Symposium for the invitation andthe financial support.

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