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The nematode pharynx has been used widely as the basis for the taxonomy of these animals,
because nuclei, organized in well-defined and constant arrays were easily visualized by light
microscopists. If the distribution of nuclei in C. elegans displayed
in figure 14 is compared with the charts for Rhabditis worked out by Chitwood & Chitwood (1938), then one sees the basic
plan is the same. However, in the present work several of the nuclei in the corpus nerve cords
were found to be epithelial or muscle cell bodies, rather than neurones, as they had been assigned
The electron microscope has shed some further light on the question of the syncytial nature of the organ. As reviewed by Bird (1971), early workers had failed to find cell membranes in the pharynx, and more recently Reger (1966) described Ascaris as having a syncytial myoepithelium in the pharynx. This complete reconstruction of the pharynx and other electron microscopic studies (Lee 1968; Yuen 1968b) clearly demonstrate that cell membranes can be observed in many species. However, it is worth noting that muscle cells in C. elegans are binucleate within sectors, dividing up generally into layers of three cells. In some places where cells are free to cross such boundaries as the marginal cells, the cells do fuse. This is seen on the ventral side of muscles m5 and for the dorsal and right g1 gland cells. Why in this last instance only the right side should become fused is unclear, but this was observed in the three animals examined. This pattern of fusion contrasts with observations on other species for which Yuen (1968a) found the subventral glands of Ditylenchus were fused in the anterior half of the terminal bulb, while much earlier Looss had described the fusion of all three gland cells of Ancyclostoma (Bird 1971). One is led to the conclusion that there is a tendency towards a syncytium in the pharynx of nematodes. In contrast, in the body of C. elegans only the hypodermal cells appear to behave similarly (White et al. 1976).
Structurally body wall and pharyngeal muscle are different. Body muscles run longitudinally
and are 'obliquely striated' (Rosenbluth 1965), while each pharyngeal muscle sector is composed of a single sarcomere running between basement membrane and cuticle. Biochemical
differences in myosin heavy chains, as well as the isolation of mutants affecting only the body
muscle myosin further distinguishes these two muscle classes (Epstein, Waterston & Brenner 1974).
Neuromuscular relations in the pharynx also differ from the somatic system. As described in the accompanying paper (White et al. 1976), the body muscles have muscle arms that extend to the nerve cords to receive synapses. In the pharynx neurones run within muscle sectors and most synapses to muscle are made en passant. Motor neurones M3 are unusual in this regard, sending a branch to the most posterior projection of muscles m4 to make their synapses.
The pharynx is a three part structure. In contrast, the pharyngeal neurones are paired subventral cells or they are single dorsal cells, often with two equivalent branches. In the case of motor neurones, paired subventral cells (M2, M3) form muscle synapses from the subventral nerve cords and then run around to the dorsal muscle sector where both cells synapse onto the dorsal muscle cell. Single dorsal motor neurones, such as M4 and M5, have one branch in each subventral nerve cord synapsing onto muscles; both branches then travel to the dorsal side to synapse onto the dorsal muscle sector. Thus, by having the dorsal sector receive synapses from both paired subventral cells or both branches of dorsal cells, the nervous system can innervate the three muscle sectors of the pharynx. In a similar fashion, the marginal cell neurones each synapse onto one subdorsal marginal cell and both neurones synapse onto the ventral cell. Also, the paired subventral interneurones synapse onto target cells on the subventral side of the nerve ring and onto the same cell and/or its partner of paired target cells on the dorsal side. Thus, the neurone classes in the pharynx are bilaterally symmetric about the dorsoventral midline. For symmetry in the single dorsal cells with two branches a line must be drawn through the cell body, so that each branch of the cell plus one-half nucleus behaves similarly to one of the paired subventral neurones. Possibly, this might suggest that these single dorsal cells are tetraploid.
Termination of neuronal growth in the pharynx may occur by fibres meeting and forming
gap junctions. Thus, when pairs of cells or branches of a single cell end on the dorsal midline,
they do so by meeting in gap junctions to each other. In the preceding paper, similar behaviour
occurring between classes of ventral cord neurones is described (White et al. 1976).
Motor neurones in the pharynx specifically innervate particular classes of target cell. For example, the motor neurones M3 make motor synapses on the posterior projection of m4. Here the neurone lies between two muscle cells m4 and m5, but synaptic bars on the m5 side of the neurone are never seen. The one exceptional animal in which several changes were seen in the nervous system provides another example of the preference neurones have for certain target cell classes. When the marginal cell neurone failed to synapse on its normal target, the ventral marginal cell, it continued to grow around the nerve ring until it eventually synapsed on another marginal cell, rather than on muscles or neurones.
Paths of cells in the nerve ring might be laid down by other cells. This is suggested from the reconstruction of the one exceptional animal. When a pair of cells that lie one on top of the other in the nerve ring were both found to have altered morphology in the ring it indicated that one cell (I4) could take its growth cues from the other (MI) and follow along this fibre, possibly recognizing a membrane polarity which leads to its asymmetric growth. If a range of alterations in the pharyngeal nervous system could be collected by isolation of mutants then further mechanisms for specifying neuronal growth might be deduced.
Several neurone classes in the pharynx anatomically appear to have several functions, aside
from their synaptic assignments as either motor or interneurones. Many have free subcuticular
endings suggesting that they are also mechanoreceptors. Motor neurones M3 could therefore
be sensory-motor neurones, similar to other neurones described in the somatic nervous system
of C. elegans (Ward et al. 1975). The NSM cells have been called secretory because of the large
vesicles seen in these cells and so could be not only sensory-motor neurones, but neurosecretory
cells, as well. Cells with the ultrastructure of neurosecretory neurones were also observed by
Lee (1968) in studies of the pharynx of Nippostrongylus brasiliensis. Possibly such cells in the
pharynx of C. elegans, which, because they lie at the outside edge of the pharynx, might secrete
a humoral factor into the pseudocoelom when their sensory endings detect food
in the lumen.
In addition to multi-functional neurones, the muscles m6 appear both to be contractile cells, and to lay down the cuticle in the pharynx. Vesicles are seen at the grinder side of m6 and in some animals the vesicles were filled with material that looked similar to the cuticle. It appeared that cuticular matter was being exocytosed toward the cuticle of the grinder at the time of fixation.
Multi-functional cells, in particular sensory-motor neurones, have been suggested as intermediates in the evolution of nervous systems (Horridge 1968 ; Coggeshall 1971). Thus, the pharynx of C. elegans appears to be not only a simple neuromuscular system, but also a primitive one.
By using Nomarski optics in the light microscope, the movements of individual pharyngeal
muscle layers can be observed in animals feeding on bacteria. When observations were made
in this way on feeding C. elegans, it was found that the description of feeding given by Doncaster
(1962) for Rhabditis oxycerca and Pelodera lambdiensis also applied to C. elegans. Briefly, in each
cycle of pumping the grinder is set in motion first, when the 'flaps' are pulled backwards by
muscle contraction in the terminal bulb causing food in front of the grinder to be processed
through. Then the corpus opens, as well as the anterior part of the isthmus, and food particles
and liquid are sucked in. This region closes again before the contraction stroke of the terminal
bulb has progressed to the point of causing ground food to pass in to the gut. After this, the grinder
flaps return to the resting, T-shaped configuration and there is a slight pause before they start
to move forward again, initiating a new cycle of the pump. Particulate food is trapped by the
corpus, while excess water is expelled when the pharynx closes. Food plugs accumulate in the
anterior half of the isthmus, and then portions are passed backwards through the isthmus to
lie just in front of the grinder.
The nematode feeds almost constantly during its life cycle, except during the moult, or when a special, resistant form, the dauer larva, is induced upon starvation. At these times a mechanism for inhibiting pharyngeal pumping is required. Furthermore, during the end of the moult, the corpus alone must be inhibited while at least the back half of the pharynx is activated for ingestion of the shed cuticle.
It seems likely that there are two different pathways for controlling pumping and its inhibition. The diagrams in figure 32 indicate a possible mechanism in which it is suggested that I1 mediate inhibition, since these cells receive input from the somatic nervous system. Neurones I2, which have connections to all motor neurones would then control pumping.
Inteneurones I2 are not connected to the somatic nervous system, and therefore each cycle
of the pump must be regenerated from within the pharynx itself. The I2 cells have proprioreceptive-like endings attached to muscle m1 and could excite M1 and initiate opening of the procorpus when it is closed. This would start a cycle of the pump. At the same time as the
procorpus opens, the metacorpus and front of the isthmus would be opened by I2 exciting M2
to cause contraction of muscles m4 and m5.
However, motor neurones M2 make synapses along the length of the single isthmus muscle m5, suggesting that the whole of this muscle should contract in response to M2, but during normal pumping only the anterior part opens. An inhibitory motor neurone in the isthmus, acting antagonistically to M2 might modulate the contraction of m5. Motor neurone M4 appears anatomically ideally suited to this. Examination of the distribution of motor synapses from M4 reveals that at the anterior of the isthmus the branches of M4 are dendritic while motor synapses are concentrated posteriorly in the isthmus. Here also, all three muscle sectors are innervated by the two subventral and two dorsal branches making muscle synapses (figure 28). Thus, inhibition from M4 would be concentrated at the back of muscles m5, and if in this region inhibition is greater than the excitation from M2, then only the anterior portion of this muscle would contract. Thus, together M1, M2 and M4 control the contraction stroke of the corpus and isthmus. This results in the intake of food particles and their accumulation in the front of the isthmus. Since the NSM have proprioreceptive-like endings in the metacorpus, they could be detectors of the food plugs accumulating here. Then by changing the tone of the isthmus muscles, the NSM might regulate the flow of food to the terminal bulb. Once food arrives in the terminal bulb, it could be detected by the subcuticular ending of I6, which would stimulate M5 to begin the muscle contractions which move the grinder. This response to food could be inhibited by the I2 connection to I6, so that this contraction in the terminal bulb never occurs simultaneously with the opening of the corpus. The initial contractions in the terminal bulb might stimulate I5 which has its possible mechanoreceptors attached to the cell bodies of M2. This interneurone might then augment the I6 input to M5 leading to further contraction of the terminal bulb muscles and the passage of food into the gut.
Somatic sensory input to the pharynx is primarily through a set of anterior sensilla, the inner
labials. These are composed of a mechanoreceptor and a possible chemoreceptor
(Ward et al. 1975). This last drives the pharyngeal somatic interneurone most directly. Thus, in response to
the outside environment the somatic interneurones may inhibit the procorpus directly and the
rest of the pharynx by way of I1. Only I6 is not connected to this circuit and could therefore
still respond to mechanical stimulation. During moulting when the cuticle is shed and ingested,
muscles in the isthmus and terminal bulb often operate while the rest of the pharynx is
quiescent. These contractions may be possible because the I6-M5 circuit is not connected to
I1 and so is not inhibited at this time.
The I1 circuit might also function in emergencies. For example, if I2 inhibit contractions in the posterior part of the pharynx when muscles m1 are contracted, then should the nematode be greedy and completely fill the corpus with a solid plug of bacteria, the I2 pumping sequence could not proceed. When this happens to feeding animals, they recover by making jerky contractions along the whole length of the isthmus, while grinding proceeds more or less normally in the terminal bulb. To act as a rescue mechanism, the proprioreceptive endings of I1 would have to detect the open state of the pharynx and by means of the connection to M2 stimulate the isthmus to contract and pass food back to the terminal bulb. Once the corpus is cleared of the food plug, normal pumping directed by I2 could resume.
Such speculations are examples of how one can combine visual observations of behaviour with the anatomical reconstruction and then suggest how a simple nervous system might work. More detailed explanations can be offered, and it is likely that from further observations of feeding animals more examples of behaviour will be collected to refine these hypotheses.
Fig. 32 - Circuit diagrams. Features of the neuronal connectivity in figures 28-30 which are seen on more than one series have been summarized in two circuit diagrams. (a) Control circuit mediated by I1. (b) Pumping circuit mediated by I2. , Motor neurone; , interneurone; , other neurones; , neurones with possible mechano-receptive ending; , chemical synapse; , gap junction; , from motor neurones point to the muscle regions of the pharynx that these cells innervate. Muscle boundaries are shown in the drawing of the pharynx.
Web adaptation, Thomas Boulin, for Wormatlas, 2002, 2003