the journal of comparative neurology volume 116 issue 2 1961 [doi 10.1002%2fcne.901160209] c. a. g....

On the Functional Anatomy of Neuronal Units in the Abdominal Cord of the Crayfish, Procambarus clarkii (Girard)

C. A. G. WIERSMA AND G. M. HUGHES Biology Division, California Institute of Technology, Pasadena, CaliforniaZ

In a previous publication (Wiersma, '58) it was shown that a number of the neurons whose axons form the so-called circumesophageal commissures of the crayfish can be characterized by (a) the nature of the sensory stimulation to which they respond, (b) the type of response, which distinguishes primary sensory fibers and interneurons, ( c ) their location in the cross section and (d) the size of their action potentials, which is related to their diameter. Using these criteria it was possible to establish 100 physiological entities, even though only two main types of sensory stimulation were used, namely touch of hairs on the body surface and manipulation of joints. The same methods have now been applied to the connectives of the abdominal cord, in which the number of fibers is about half that in the commissure.

A report has already been presented of the types of physiological connections made by axons of the cord, without entering into their individual properties (Hughes and Wiersma, '60a). The purpose of the present paper is to classify the data obtained for the cord in a manner comparable to that for the commissure. Only those entities are described ( A 1 to A 75) which have been repeatedly found in a number of preparations. They are presented in table 1, to which the reader is referred for all those cases in which only the number of the entity is used in the text. The results were collected from about 120 preparations, the great majority of these being isolated abdomens.

METHODS

The methods were similar to those used for the analysis of units in the crayfish

commissure, and have been described in detail elsewhere (Hughes and Wiersma, '60a). It was found that the isolated abdomen preparation had considerable ad- vantages over that of the whole animal. Its survival time was at least as long, and in general it remained in better condition than whole animal preparations. The best results with the latter were obtained when, in addition to exposure of the cord, the circumesophageal commissures were also prepared. It is likely that this difference was due to improved circulation; replace- ment of blood by the perfusion solution (van Harreveld, '36) is greatly facilitated by opening the thoracic cavity. It may be that clotting of blood is one of the factors producing untimely deterioration of the preparations.

The cord was exposed from the ventral side by stripping off a median part of the sternal area between the second and 6th abdominal segments. Great care was taken to keep all roots of the ganglia in- tact. First roots are especially easily damaged, and inspection and observation of spontaneous movements of the swimmerets was routine in order to exclude this possibility.

For several reasons splitting of the cord was a source of possible experimental error. The surrounding membrane is very tough and it takes considerable force to penetrate it. This probably leads to dam- age of a number of axons in each preparation along the line of split, which was usually in the horizontal plane, dividing the cord into about equal parts. The next

'On leave of absence from Zoological Depart-

ZSupported by grant G-5461 of the National ment, Cambridge University, England.

Science Foundation.

209

210 C. A. G. WIERSMA AND G. M. HUGHES

TABLE 1 Short descriptions of all fibers running in the connective between the third and fourth

abdominal ganglion discussed in the text, are listed in numerical order The location of the sensory fields to which the fiber responds is given after the number,

followed by the type of stimulation to which it responds. Next is indicated whether the response is obtained from the same side of the animal as the prepared connective (Horn.), from the other side (Het.), from both sides (Bil.), or that the response is asymmetrical (As.). L. refers to location in cross section as presented in figure 1. Id. C. indicates that the fiber is identical in reaction with the commissure fiber with that number (Wiersrna, '58).

A 1. A 2. A 3. A 4. A 5. A 6. A 7. A 8. A 9. A 10. A 11. A 12. A 13. A 14. A 15. A 16. A 17. A 18. A 19. A 20. A 21. A 22. A 23. A 24. A 25. A 26. A 27. A 28. A 29. A 30. A 31. A 32. A 33. A 34. A 35. A 36. A 37. A 38. A 39. A 40. A 41. A 42. A 43. A 44. A 45. A 46. A 47.

Abbreviations used in table App., appendages Dors., dorsal Abd., abdominal Vent., ventral Seg., segment SRI, tonic stretch receptor Med., medial location of the interneuron SRB, phasic stretch receptor in the cord Sw., swirnrneret Lat., lateral location of the interneuron Tels., telson in the cord Urop., uropod Pl.pl.1 or II., pleural plate hairs innervated U. Seg., uropod segment (6th abdominal)

Prim., bundle of primary sensory fibers

by the first or second root

Abd. Seg. Joint. Tonic. Extension. Horn? L78 SRI-Ab 5/6. Horn. L78-79 Urop., Tels. Hair. Horn. L82

Id. C 100 Id. C 19

SRI-Ab 4/5. Horn. L78 Sw. 2-Urop. Basal Joint. As. L82 Urop., Tels. Hair Fringe. Prosser's. Horn. L85 Abd. Seg. 4-U. Seg. Hair. Horn. L82 SRI-Ab 6/T. Horn. L78 Abd. Seg. 5. Hair. Prim. L84 Sw. 1-Urop. Basal Joint. Phasic. Horn. L84 Sw., Pl.pl.1. Seg. 1-5 Hair. Bil. L84 Abd. Seg. 3-5. Hair. Horn. L83 Pl.pl.1. Seg. 2-5. Hair. Horn. L85 Abd. Seg. 5. Hair. Med. Horn. L78 U. Seg. Hair. Med. Horn. L78 Abd. Seg. 3. Hair. Prim. Horn. L83 Tels. Hair. Horn. L83 Abd. Seg. 4. Hair. Prim. Horn. L84 Sw., Pl.pl.1. Seg. 2-5. Hair. As. L80 Urop. Joint. Tonic. Horn. L84 SR2-Ab 5/6. Horn. L79 Abd. Seg. 4. Hair. Het. L79 Abd. Seg. Joint. Phasic. Extension. Horn? L79 Sw. 1-Urop. Joint. Phasic. Bil. L81 SR1-Ab 3/4. Horn. L79 Sw. 4,5. Joint. Prim. Horn. L83 Sw. 2-5. Basal Joint. Horn. L83 Sw., Pl.pl.1. Seg. 2-U. Seg. Hair. Horn. L84 Abd. Seg. 2. Hair. Het. L82 Abd. Seg. 3. Hair. Het. L82 Sw. 4. Basal Joint. Tonic. Horn. L81 Urop., U. Seg., Tels. Hair. Horn. L80 Sw., Pl.pl.1. Seg. 3 Hair. Joints? Horn. L81 Abd. Seg. 2,3. Hair. Het. L84 Sw. 5. Basal Joint. Tonic. Horn. L81 U. Seg. Hair. Quack. Horn. L81 Urop., Tels. Hair. Dors. Vent. Het. L81 Abd. Seg. 3,4. Hair. Horn. L79 Abd. Seg. 4. Hair. Med. Horn. L79 Abd. Seg. 2,3. Hair. Horn. L83 Abd. Seg. 2-5. Hair. Horn. L79 Abd. Seg. 3. Hair. Med. Horn. L79 Anal Valve. Joint. Horn. L82 Sw. 4. Joint. Prim. Horn. L78 Sw. 5. Joint. Prim. Horn. L78 Abd. Seg. 1-U. Seg. Hair. Dors. Vent. Horn. L83 Sw. 3. Joint. Prim. Horn. L83

- . ~ _ .

Id. C 89 Id. C 48

Id. C 91

Id. C 65 Id. C 38

Id. C 15

Id. C 93

Id. C 3

Id. C 71

CRAYFISH ABDOMINAL CORD NEURONS 21 1

TABLE I-(Continued) Sw. 3. Basal Joint. Prim. Tonic. Horn. L76 Abd. Seg. 5. Hair. Lat. Horn. L80 Abd. Seg. 4. Hair. Lat. Horn. L80 Pl.pl.1. Seg. 3. Hair. Horn. L84 Abd. Seg. 2-5, U. Seg.? Hair. Dors. Vent. Bil. L78 SRI-Ab 2/3. Horn. L78 Abd. Seg. 2-5, U. Seg.? Hair. Het. L78 Sw., Pl.pl.1. Seg. 2,3. Hair. Horn. L84 U. Seg. Hair. Lat. Horn. L80 Abd. Seg. 5, U. Seg. Hair. Horn. L80 Abd. Seg. 4,5. Hair. Horn. L85 Urop., Tels. Hair. Vent. Horn. L77 U. Seg. -5th. Joint. Tonic. Extension. Horn? L81 SRI. All Seg. Joint Tonic. Horn. L77 Abd. Seg. 3. Hair. Lat. Horn. L85 Abd. Seg. 2-4. Hair. Horn. L84 Abd. Seg. 2-U. Seg., Tels? Hair. Bil. L84 Abd. Seg. 5. Hair. Het. L78 Sw., Pl.pI.1. Seg. 3. Hair. Joint. Het. L76 Sw. 3. Hair. Horn. L76 Urop. Basal Joint. Horn. L77 Urop. Basal Joint. Het. L77 Urop. Joint. Bil. L78 Abd. Seg. 3-5. Hair. Bil. L80 Anal Valve. Joint. Het. L76 Abd. Seg. 4- U. Seg., Urop., Tels. Hair. Horn. L83 Sw., Pl.pl.1. Seg. 3. Hair. Prim. Horn. L83 Sw., Pl.pl.1, Seg. 5, Hair. Joint. Horn. L84

A 48. A 49. A 50. A 51. A 52. A 53. A 54. A 55. A 56. A 57. A 58. A 59. A 60. A 61. A 62. A 63. A 64. A 65. A 66. A 67. A 68. A 69. A 70. A 71. A 72. A 73. A 74. A 75.

Id. C 51

Id. C 45

Id. C 56

step was to peel the membrane surrounding one of the connectives, usually the left, though sometimes the right or both connectives were used. In general no active fibers clung to this membrane but fibers adjoining the membrane may well have been damaged during peeling.

Figure 1 shows the areas into which the cross section of the connectives was divided in order to determine the locations of the small bundles into which it was split during an experiment. The accuracy of this procedure was greater than for the commissures since, in contrast to the commissures, the shape of the abdominal connectives shows little variability between preparations, which considerably reduces a major source of error in localization. However, another source of error, especially in the earlier experiments, was the ease with which rotation of the major bundles occurred once the sheath around the cord had been removed. Localization became much more precise after a number of marker fibers had become established.

Most preparations were of the connectives between the third and 4th ganglia, but in a large number those between the second and third ganglia were used, and

Primary sensory fibers present differ mark- edly in the different connectives, whereas on the other hand few interneurons are found in only one of these places and not also in the others. All numbers used in this paper refer to fibers present in the cord between the third and 4th ganglia.

Determination of the properties of single entities was simpler than in the commissure, both because of a greater pre-

Fig. 1 Cross section of right connective between third and 4th abdominal ganglion with the numbers of the areas into which it was divided. Large structures on top are the medial (76) and lateral giant fibers. some were made between the 4th and 5th.


ponderance of fibers with simple reactions and the large reduction in the extent of the total sensory area which needed to be explored. As a result the number of units found in one preparation was generally much higher than that in similar commissure experiments. Of the total 75 entities established, as many as 30 have been rec- ognized in a single preparation, and it was routine to find more than 15. Hence the number of times individual units were found in the cord is in general higher than in the commissure, although the total number of preparations was considerably smaller (about 1/5).

RESULTS

Primary sensory fibers in the abdominal connectives form a much larger part of the total number of axons than in the commissures. In both parts of the central nervous system the fibers from a common source and with similar function run together in bundles, With the exception of the primary sensory fibers of the abdominal stretch receptors, which can be prepared as single units, a single number (A 9 etc.) has been given to such bundles of primary sensory fibers. Both ascending and descending bundles are found.

Primary sensory fibers responding to touch of hairs on each abdominal segment enter through both first and second roots and will be divided into groups according to this difference (see fig. 2).

Primary sensory hair fibers f rom first root innemation (figs. 3, 4)

A . Ascending. None found. B. Descendin.g. A 74. Primary sensory

fibers from hairs on the third swimmeret and posterior pleural plate ridge and anterior 4th pleural plate ridge, which are known to enter through the first root of the third ganglion, form a bundle in area 83. This bundle consists of a relatively large number of small fibers. In the lead between second and third ganglia a similar bundle of descending fibers from the second ganglion replaces these fibers. This bundle was called A 74 ( 2 / 3 ) . A 74 was found 4 times, A 74 (2/3) 4 times.

The primary hair fibers entering through the first root appear to have a restricted distribution, compared to other

Summary.

PLEURAL PLATE , SEGMENT 4 - - ? + / TERGUM

m d i c q m THIRD GANGLION

Fig. 2 Schematic representation of the third and 4th abdominal segments, showing the areas innervated by the first and second root of the right half of the third abdominal ganglion. Dark shading, first root and areas innervated by it, light shading second root and its areas. Arrow shows the location of the abdominal stretch receptors, excited by bending the 5th on the 4th segment, innervated by the second root of the third ganglion. (From Hughes and Wiersma, '60a)

sensory fibers. Like all others they are restricted to the homolateral connective, but run in this only in a caudal direction for the length of a single interganglionic space. There seems to be no histological evidence for fibers with such a course. From their function in local reflexes it appears likely that they synapse with other fibers both in the ganglion they enter and in the one more posterior.

Primary sensory hair fibers from second root innervation (figs. 3, 4)

A. Ascending. A 9. Primary sensory fibers of 5th abdominal segment. There are a number of these small fibers, which react separately to touch of small areas. These fibers have also been found several times between second and third ganglia. Though they may all enter through the second root of the 4th ganglion, they must run well forward. Location 84, near 83. Found 20 times, is used for marking the area.

213 CRAYFISH ABDOMINAL CORD NEURONS

A 1 8 / I \ A 47 A 1 6

Fig. 3 Cross section showing the location of the bundles of primary sensory fibers, responding to touch of hairs and to joint movements of swimmerets. Slight differences of location between diagrams and text (e.g. A 48) were prompted by considering neighboring fibers in the former.

A 18. Primary sensory fibers of 4th abdominal segment. Again there are many small fibers in a bundle, which runs near the A 9 bundle. In the lead between the second and third ganglia only ascending fib-

B . Ascending a n d descending.

ers are found. Location 84. Found 24 times, often with A 9 and A 16.

C . Descending. A 16. Primary sensory fibers of the third abdominal segment. In the connective between second and third ganglia they are present as both small ascending and descending fibers. They were not found in 4/5 leads. Location 83 close to 84. Found 15 times, of which about half in second to third connective.

No primary fibers descending from the hairs of the second segment were found in the connective between third and 4th ganglion, though they occur in 2/3 leads.

In contrast to the first root hair fibers, those of the second root, representing hairs on the dorsal surface, run much further through the cord. They de- scend only for a single segment, and these branches may well be direct continuations of the peripheral fibers, since Retzius (1890) has noticed a number of fibers entering through the second roots, which bend immediately backward, without connections in the ganglion. Their forward course can certainly span three interganglionic distances, as in the case of fibers from the 5th segment found in 2 / 3 leads. Remarkably, the dorsal hair fibers of the uropod and telson segments appear to

Summary.

9

74 Fig. 4 Schematic drawing of side view of abdomen indicating areas supplied by bundles of

primary hair fibers.


have a much more restricted course. They have not been encountered even in 4/5 leads, except in a single instance. This is in sharp contrast to the ease with which all other bundles were found, so that, un- less more evidence is obtained, their regular presence cannot be accepted. If so it might indicate that the second root hair fibers of the anterior abdominal segments are separately integrated from those of the posterior region, possibly in the first abdominal or last thoracic ganglion.

Primary proprioceptor fibers entering through the first root (fig. 3)

The decision whether a joint fiber is primary sensory or an interneuron is much more difficult to make than for hair fibers. In the following enumeration there may be some errors made in this respect. One criterion used, is that fibers forming very clear bundles, whose responses correspond with that obtained when a peripheral lead from the root itself was taken, were considered as primary. In the cord other proprioceptor fibers are more solitary and are sometimes of considerable size. When they responded to the movement of what appeared to be a single joint of a single swimmeret, they were considered as primary, otherwise as interneurons.

A. Ascending. A 26 and A 26A. A bundle of small primary sensory fibers from the 4th and 5th swimmerets. These are always close together and in most cases could not be split. Different types of fiber are found, tonic and phasic and separate fibers for different joints. For instance, a certain fiber in this bundle reacts on spreading of the exo- and endopodite. In all these respects the bundle gives the same kind of signals as can be obtained with a peripheral lead from the first root of these ganglia. These fibers may well run for shorter distances in the cord than the following two entities. For in contrast to the latter, there were indi- cation that only a few if any of the fibers from the 5th swimmeret also appear in 213 leads. This bundle was found 19 times in area 82, near 83.

A 44. Primary sensory fibers of joints of the 4th swimmeret. These form a much smaller bundle of medium-small fibers in area 78. They respond solely to backward

movement of the swimmeret and are tonic. They might well be fibers connected with the muscle receptors of Alexandrowicz ('58), if such are present for the swimmerets, for they appear to react only during active contraction. However, it is not completely excluded that they are interneurons. The responses can be inhibited by moving the 5th swimmeret forward and excited by the forward movement of the second swimmeret. They undoubtedly play a part in the generation of the metachronal rhythm of the swimmerts. They are located dorsal of A 26 in area 78 and one or more of them were found 12 times.

A 45 appears to be a similar small fiber for the 5th swimmeret which fires tonically, especially during active backward contraction of the swimmeret. It can become inactive although the swimmeret maintains its position passively, again indicating that a muscle receptor organ may cause the firing. It is a close neighbor of A 44, running like the latter near the abdominal stretch receptor axons in area 78. It was found 7 times.

B . Descending. A 47. Homolateral third swimmeret, joint fibers both tonic and phasic, different fiber sizes. These fibers were found in the same area in 2/3 leads, where they are ascending. Location 83 near 82. Found 10 times.

A 48 resembles A 44 and A 45, giving a tonic discharge, especially during backward contraction of the swimmeret. As in the case of A 44 more than one fiber was occasionally found being medium and small in size. Location 76. Found 6 times, three of them in 2/3 leads (ascending).

A 47 and A 48 are certainly different entities, as indicated by their locations, and by the fact that they have been found in pairs in single preparations.

Summary. The absence from 3/4 leads of fibers from the second swimmeret may indicate that this type of fiber descends only one connective, whereas the presence of fibers from all swimmerets in 2/3 leads shows that they ascend for at least three successive connectives. If the same held for the uropod joint fibers, one would ex- pect to find them as readily in the 3/4 lead, but this was not the case. Reactions in single axons to uropod joints were found in several instances, and although

CRAYFISH ABDOMIN 'AL CORD NEURONS 215

it is uncertain whether these are definitely interneurons, they will be regarded as such in this paper. Two other fibers specific for a single swimmeret have been found (A 31 and A 35) which might be- long here, but which will be described under interneurons.

Primary proprioceptor fibers entering through the second roots (fig. 5)

Whereas previously (Wiersma, '58) the neurons of the abdominal stretch receptors were named RM1 for the slowly adapting and RM2 for the fasting adapting type, the more accurate designation of SR1 and SR, (Wiersma and Pilgrim, '61) will be used in this paper.

A. Ascending. A 2. Tonic stretch receptor of the joint between 5th and uropod (6th) segments (SRI-Ab 5/6). This fiber enters through the second root of the 4th ganglion. The fiber runs on to the brain and is C. 100 of the commissure. It is large and located as are all other abdominal stretch receptor fibers in areas 78 and 79 under the medial giant fiber. Found very many (21 recorded) times. A 21. Phasic stretch receptor SR,Ab 5/6. This fiber is even larger than A 2. Location 79, found 11 times.

A 8. Tonic stretch receptor of telson on uropod segment (SR1-Ab 6/T). This is a smaller fiber, which has so far not been

A 21 I

A 6

A4 Fig. 5 Cross section with location of the

primary sensory fibers of the stretch receptors of the abdomen and those s o n s responding to extension of abodminal segments.

found in the commissure. It enters, with its partner the phasic receptor, through the homolateral second root of the 5th ganglion. Found 13 times, location 78.

Because the phasic receptor of this joint was found ony twice, it has not been given a number, but it was obtained in a 2/3 lead, indicating that all these primary sensory fibers go to the brain.

B . Descending. A 4. SRI-Ab 4/5, entering the cord through second root of third ganglion. This large fiber is ascending in 2/3 leads, and is identical with C89. Again it is accompanied by the fiber of the phasic stretch receptor of this joint, which was found three times. Location 78, found 11 times.

A 25. SRI-Ab 3/4. Found 13 times. It is large and is identical with C93. Loca- tion 78.

A 53. SRl-Ab 2/3 is identical with C51 of the commissure. Found 6 times. Loca- tion 78. The descending branch appears to be medium in size.

Because they are also most likely primary fibers, though this is not strictly proven, some fibers which react to extension of the tail, but which do not enter the cord through any of the roots of the ganglia, and appear to be excited within the cord will be next discussed. The reasons for ascribing extensor sensation as the function of these axons has been given elsewhere (Hughes and Wiersma, '60a).

A 1. Tonic extensor fiber for at least the posterior joints and perhaps all joints of the abdomen, but not of the one from uropod segment to telson. Details of the responses of this fiber are given in the paper quoted (Hughes and Wiersma, '60a). This fiber has been found many (16) times. Location: area 78. There are in reality two tonic fibers, a smaller and a somewhat larger one, which run together with the phasic fiber (A 23). The smaller one is more tonic but stops when extension is too extreme. They resemble each other so closely that it is not possible to determine which one has been found when a single signal is obtained.

A 23. Phasic extensor fiber of the abdomen. It is larger than the previous ones, and located near them and the stretch receptor fibers. Found 15 times in area 78-79.


A 60. Also reacts on extension, but for this fiber it has not been proven that the sense organs are not innervated by a root. It is more restricted in its sensitivity, an- swering with low frequency exclusively to extension of the joint between 5th and uropod segments but quite tonically. Its location is rather removed from that of the previous fibers, namely area 81. It was found 6 times. It is a medium-sized fiber, found also in preparations in which A1 was present. This fiber may well be an interneuron.

Summary. The finding of SR2 fibers in the cord makes it likely that all of the quick stretch receptor cells also send direct branches to the brain. This was expected from results in the commissures (Wiersma, 58). According to the histological find- ings (Allen, 1894; Alexandrowicz, 51) these fibers should also have descending branches, and there are two technical reasons for the failure to find their signals in the present experiments. The anterior SR,s are known to adapt quicker than the more posterior ones, and quick bending of the anterior joints is very difficult to obtain in the isolated abdominal preparation as used. The fact that the extensor fibers run in the same bundle as the SR fibers enhances the likelihood that integration of all these signals takes place both in the brain and in the 6th abdominal ganglion. Such integration, of which the main function may be the regulation of the activity of the central giant fibers, is indicated by the existence of fiber A 61 (see below).

Interneurons The distinction made above between as-

cending and descending fibers, was not altogether appropriate for those of the stretch receptors and perhaps other primary sensory fibers and can lead to con- fusion when applied to interneurons. Only in some cases is there no objection to this term, as for example the interneuron col- lecting from hairs on the telson is ascending throughout the central nervous system. But the term is quite inappropriate when the interneuron collects impulses in both the third and 4th ganglia so that impulses pass in both directions in the connective between these ganglia (Hughes and Wiersma, 60a). Because relatively

few interneurons are excited by sensory fibers which enter through both first and second roots of abdominal ganglia 1-5, these terms retain value for descriptive purposes. However, the fact that the roots of the uropod and telson ganglion (6th) cannot readily be homologized with first and second roots, diminishes their appro- priateness.

As in the commissure, interneurons can readily be divided into homolateral, heterolateral, bilateral and asymmetric depend- ing on whether the sensory field to which they respond occur on the same side of the body as the connective in which they run, on the other side, on both sides symmetrically, or on both sides but with an asymmetrical distribution of their sensory fields. They can also be distinguished according to the type or types of sensory input to which they react.

It should be realized that there are certainly a number of interneurons present in the cord which receive their sole input in the ganglia of the thorax and the brain. These were encountered in whole animal preparations but have not been found enough times to be described here. It will be noticed from the diagrams showing the distribution of the axons found, that fewer are present in areas 76-77 than would be expected from a random distribution. There is strong reason to believe that a considerable number of descending interneurons, but by no means all, are located in these dorsal areas underneath the giant fibers.

The order followed in describing interneurons will be to start with those responsive to hairs, next those responding to proprioceptive stimulation and finally those activated by both these methods. First, the interneurons stimulated by hairs on the dorsal aspect of the abdomen, will be discussed, first all those with sensory fields on one homolateral segment then on one heterolateral segment, next the same for two segments and then three and more segments. The interneurons responding to the ventral aspect will be similarly treated, followed by those responding to both aspects.

Interneurons stimulated by hairs o n dorsal areas. The remarkable distribution of the nerves on the dorsal surface of

CRAYFISH ABDOMINAL CORD NEURONS 217

each anatomical segment (which for all abdominal segments, including the uropod (6th) segment are innervated by roots of two different ganglia, fig. 2) makes it uncertain in most of the following cases whether the interneurons respond to the external anatomical or the neurological segmentation or to neither exactly. When an interneuron is described as reacting to, e.g., 4th and 5th dorsal abdominal segment, it means that it reacts to dorsal stimulation of both segments but that reaction to the posterior part of the pleural plate 11 innervation of the third segment, innervated by the second root of the third abdominal ganglion, is not excluded. It will be clear from the following that interneurons can also react to only a part of these sensory fields. Exact mapping is very difficult, either because of the neces- sity of stimulating a large area in order to make the interneuron fire or on account of its extreme sensitivity to a few hairs, and has therefore not yet been tried in many cases.

Interneurons from dorsal areas o f single abdominal segments (figs. 6, 7)

A. Homolateral interneurons. For each segment two fibers have been found. They differ, as fa r as is known only in their locations within the cord. There appears to be distinct bundle formation, one bundle running near the abdominal stretch receptor

A I4 Fig. 6 Cross section with the location of inter-

neurons responding to touch of hairs of dorsal side of abdominal segments; 0, homolateral interneurons; (>, heterolateral.

HOMOLATERAL HETEROLATERAL

29

30

22

65

I7 Fig. 7 Dorsal view of abdomen with areas

innervated by interneurons which cover only small dorsal parts, limited to single segments.

fibers, and thus in the medial half, the other bundle being more laterally located. All these interneurons appear to react to the sensory input of a single root, and thus to the posterior pleural plate 11 area of one segment and the dorsal area of the next (fig. 2).

A 42. Medial interneuron for homolateral dorsal third segment hairs. Me- dium sized fiber located in area 78-79. Similar to A 14, A 15 and A 39, of which it is a neighbor. It was found 11 times.

A 62. Lateral interneuron for dorsal third segment hairs. Homolateral, medium in size, and located in area 85. Like the previous fiber, it is found only in diphasic and anterior recordings. Found 6 times.

A 39. Medial interneuron for dorsal 4th segment homolateral hairs. It is a neighbor of A 14 and A 15 in area 78-79. Large, it was found 10 times. This fiber can be obtained in both anterior and posterior recordings, which makes it necessary that the primary sensory fibers make synaptic connection with it in both the third and the 4th ganglion.

A 50. Lateral interneuron of the 4th segment active in anterior and posterior recordings. Medium in size, it is a neighbor of, and similar in response to, A 49 and


A 56. Found 6 times, and located in area 80.

A 14. Medial interneuron for the dorsal 5th abdominal segment hairs. It is very like A 15 in its properties and location. Large, and located in area 78, it was found 20 times.

A 49. Lateral interneuron of dorsal hairs of 5th abdominal segment. Very like A 56 in properties and location in area 80. Large, it was found 6 times, once in the same preparation as A 14.

A 15. Medial interneuron of homolateral hairs on dorsal aspect of uropod segment only. A large fiber in area 78 which very closely resembles the next one (A 56). Found 21 times, it sometimes responded in addition to hairs on the basal joint of the uropod.

A 56. Lateral interneuron for homolateral hairs on uropod segment. Located in area 80, this large fiber was found 7 times, twice in the same preparation as A 15. Both A 15 and A 56 may receive their main input by way of the second root of the 5th ganglion.

A 36 (fig. 13). This very large fiber has received the nickname of quack fiber. It usually has a high threshold and fatigues rapidly. Its sensory field is somewhat uncertain, but after mild fatigue it responds only to a quick brush stroke dorsally along the base of the uropod near the 5th segment. Similar fibers, located in the same part of the cross section, have occasionally been found for more anterior segments. Its location is near the border of areas 81 and 85, it was found 12 times.

B. Heterolateral interneurons. Inter- neurons for single heterolateral dorsal segments have also been found. They form a bundle very near the midline and like the homolateral fibers, appear to respond to the sensory input of individual second roots.

A 29. Heterolateral fiber of second segment dorsal hairs. This fiber is a close neighbor of A 30 and also large. Location 78-82; it was found 9 times.

A 30. Heterolateral interneuron for dorsal hairs of segment 3. This is also obtained in 2/3 leads as is the previous fiber. Large fiber, located in area 78-82 near the other heterolateral single segment fibers. Found 10 times.

A 22. Heterolateral interneuron of the 4th dorsal segment hairs. It is a large fiber, located in area 78-79, near A 65. Found 13 times.

A 65. Heterolateral dorsal hair fiber of abdominal segment 5. Medium sized, located in area 78 and found 4 times.

Interneurons integrating dorsal hairs of two abdominal segments (figs. 8 and 9).

A 34 \ A 40

Fig. 8 Cross section with location of interneurons responding to touch of hairs on two adjacent abdominal segments.

34

3 37 Fig. 9 Dorsal view of abdomen indicating

areas innervated by interneurons covering areas of two segments.


A number of interneurons integrate the impulses from two abdominal segments. None of these nor any of the single segment interneurons were found in the commissure. (A 32 might be considered as the last of this series but its presence in the commissure as C 3 is against this view ) .

A 40. Homolateral dorsal aspect of second and third abdominal segment. Lo- cation 83, medium in size, this fiber was found 9 times.

A 38. Homolateral dorsal aspect of abdominal segments three and 4. This medium sized fiber was found 14 times. Location 79.

A 58. Homolateral dorsal aspect of 4th and 5th segments, medium in size. Loca- tion 85. Found 8 times.

A 57. Homolateral dorsal aspect of 5th and uropod segments, medium in size, location 85. It was found 8 times, close to and also in the same preparation as A 56.

A 34. This is the only heterolateral dorsal hair fiber established for two segments, namely second and third, but it might also react to segment 1. It is a medium sized fiber located in area 80 and was found 10 times.

Unlike the interneurons for single segments, these two segment interneurons do not run in bundles (fig. 8).

Interneurons reacting to three or more dorsal segment hairs (figs. 10, 11). In contrast to the single and two segment fibers, many of these have also been found in the commissure.

A 63. Homolateral dorsal hairs of segments 2, 3 and 4. Located in area 84, medium in size, and found 5 times. It is identical with C 45.

A 12. Homolateral dorsal hairs of abdominal segments 3, 4, and 5. Identical with C 65. A large fiber in area 83, it was found 16 times.

A 7. Homolateral dorsal hairs on abdominal segments 4, 5 and uropod segment, not on telson or uropod. It is identical with C 91. It is a large fiber located in area 82, and found 17 times.

A 41. Homolateral dorsal hairs of abdominal segments l?, 2-5th segment, not on telson or uropods. Medium in size, it is identical with C 71. It was found 9 times, location 79 (see fig. 12).

A 73. Homolateral dorsal hair fiber of abdominal segments 4, 5 and uropod segment, uropod and telson. Identical with C 56, medium in size, location 83. Found 4 times (see fig. 12).

In addition to these homolateral fibers, some were found with a heterolateral and a bilateral distribution :

A 54. Heterolateral dorsal hairs of all abdominal segments, 3-5 for certain but possibly also two and uropod segment.

A 71

Fig. 10 Cross section with location of interneurons covering three or more adjacent abdominal segments, dorsal hairs; 0, bilateral interneurons.

HOMOLATERAL HETEROLATERAL I l l

54

Fig. 11 Dorsal view showing areas innervated by interneurons responding to dorsal hair touch of three adjacent abdominal segments.

220 C. A. G . WIERSMA AND G . M. HUGHES

HOMOLATERAL BILATERAL

71

Fig. 12 Dorsal view showing areas innervated by interneurons responding to more than three adjacent abdominal segments.

There might be two rather similar fibers in this area. Location 78, found 9 times, mostly restricted to segments 2-5.

A 71. Bilateral interneuron for hairs on abdominal segments 3, 4 and 5. Loca- tion in area 80 and found 7 times, of medium size (see fig. 12).

A 64. Bilateral interneuron for dorsal hairs of the whole abdomen. This medium sized fiber is most likely identical with one of the several fibers in the commissure, each of which covers this area in addition to others. Location 84, found 8 times, of which in a few cases the dorsal aspect of the uropods themselves did not give a reaction.

Dorsal hairs from telson and uropods (fig. 13). The interneurons specific for these areas are separately treated because it is difficult to compare the telson with the abdominal segments and the function of the uropods is specialized. Neverthe- less some of these interneurons may be members of one of the previous series.

A 3. Homolateral dorsal hair fiber of telson and uropod, not on uropod segment. This fiber is usually found in area 82. However, in a number of cases a similar fiber was found in more lateral areas, but then never also in area 82. It seems possible, therefore, that A 3s localization is more variable than that of other interneurons. It was found 21 times, of which

4 times in the same preparation as A 6. It was located 11 times in the medial and 7 times in the lateral half of the cord and twice near the midline. Like A 6 it is a large and sensitive axon. It is identical with C 19 (see fig. 9).

A 32. Homolateral dorsal hair fiber of telson, uropod and uropod segment. Lo- cated quite consistently in area 80, near the center of the connective. This fiber cannot be confused with A 6, but could with A 3. However, they have been found twice in the same preparation. This fiber is large, and was found 11 times. It is identical in reaction to C 3 of the commissure fibers (see fig. 11 ).

A 6. This fiber may be called Prossers fiber, as it reacts typically in the way described in his experiments (Prosser, 35), which showed the relation between touch of hairs on the fringe of the telson or uropod and the firing of an interneuron. It was found to fire for all those homolateral hairs which form the fringes of the telson and the uropod, whereas in some cases but not in others, it also fired readily when the frontal edge of the anterior rim of the uropod was touched. Of high frequency discharge and often very sensitive, the fiber can also be made to fire by bending the outer flap of the endopodite of the uropod or the proximal flap of the telson, which may be due to the stretch of the primary hair fibers. It

Fig. 13 Cross section with locations of interneurons responding to hair touch on dorsal uropod and telson areas. Note that A3 is represented twice (explanation see text).

CRAYFISH ABDOMINAL CORD NEURONS 22 1

is a large axon, in area 85, found 18 times, of which some were in 2/3 leads and also 4/5 leads. Its characteristic difference from A 3 is that the latter does not readily respond to touch of the fringe hairs, but only of the dorsal hairs of these parts.

A 37. Heterolateral uropod and telson hair fiber, responds to both dorsal and ventral aspects. Of medium size, its location is area 81. It was found 16 times, in a few of which either the dorsal or the ventral side was insensitive (see fig. 9).

A 17. Responds to hairs on the middle dorsal aspect of the anterior part of the telson. It is identical with C 15 of the commissure, but responds most likely to homolateral stimulation. The fiber is rather large, located in area 83, and often gives a tonic discharge which is inhibited by touch of dorsal hairs on both uropods and uropod segment, which was also true of C 15. It was found 9 times (see fig. 7).

Summary. The outstanding feature of this section is the observed abundance of interneurons representing the dorsal abdominal hairs. The representation of these hairs by interneurons is as great as that of the claw. For instance, stimulation of dorsal hairs on the 4th abdominal segment results in the excitation of 14 interneurons in the cord as against 11 for the claw in the commissure. The presence of so many interneurons does seem to indicate a complex set of reflexes, which differ with the extent of the touch of the abdominal segments. But, besides the fact that such reactions have so far not been noticed, it seems highly unlikely that they would serve any good purpose. Therefore, the question of redundancy of information on stimulation of single areas is here especially poignant, and a further study using whole animals should throw light on this problem.

Interneurons responding to ventral areas of the abdominal segments (fig. 14 15). Compared to the abundance of the representation of the dorsal areas, few interneurons integrate the stimulation of ventral hairs.

A 55. Reacts to hairs on both pleural plate and swimmerets of segments 2 and 3. It is a small fiber located in area 85, and was found 7 times.

A 51. A large interneuron which is fired by stimulation of the posterior pleural plate hairs of the third segment and the anterior ones of the 4th. It is thus restricted to the first root neural segment. Location 84, found 9 times.

A 67. Responds only to stimulation of the hairs on the third swimmerets, not on pleural plate 1 area. This is naturally a descending fiber, which is rather small

A 59

52

Fig. 14 Cross section with locations of interneurons responding to hairs on the ventral aspects of the abdomen.

HOMOLATERAL ASYMMETRIC

19

Fig. 15 Ventral view of abdomen indicating areas for some of the interneurons responsive to touch of hairs.


and located in area 76 and found 4 times. In the same area an ascending interneuron for the 4th swimmeret hairs has been found twice.

A 59. Responds to ventral hairs on the homolateral uropod and telson surfaces. Medium in size, located in area 77; it has been found 5 times.

The absence of ascending fibers from the more posterior swimmerets may indicate that this type of information is car- ried mainly in a caudal direction. The following 4 fibers responded to stimulation of ventral hairs on more than one segment.

A 13. Homolateral pleural plate fiber of the abdominal segments. This large fiber, located in area 85, is made to fire exclusively by the hairs on the outer edge of the pleural plates. Found 20 times. It does not usually respond to the relatively few pleural hairs of the uropod segment, but in some cases did so.

A 19. Similar to A 13, but in addition responds to stimulation of the other ventral hairs, including those of the swimmerets of abdominal segments 2-5. In some instances, this medium sized fiber also responded to stimulation of the heterolateral swimmeret and pleural plate 1 area of the third segment but never to any others. It was found 20 times, on 5 of which the heterolateral pleural plate response was present. Even so it is not yet possible to decide whether there are two fibers which differ in this one respect, or whether the same fiber can have both response types. It is unlikely that the heterolateral pleural plate response is due to an additional neighboring fiber with a signal of the same strength.

A 28. A medium sized fiber which responds to the homolateral first root hair areas of abdominal segments 2-5 and in addition to hairs on the homolateral uropod, both ventral and dorsal, but not on the telson or dorsal uropod segment. Lo- cated in area 84, it was found 13 times, once in a whole preparation, when it also responded to the thoracic leg hairs. It is undoubtedly a fiber with a larger sensory area than given here.

A 11. A bilateral fiber for swimmeret and pleural plate I hairs, it responds to stimulation of all abdominal swimmerets, but not to uropods. A medium sized fiber

found 17 times in location 84. In anterior recordings it sometimes continues to respond to all of its sensory areas, in other cases only to those in front. Similarly in posterior recordings responses to the whole sensory field or only segments 4-5 may persist.

Interneurons activated by both dorsal and ventral hairs (fig. 14). Two such fibers were found, one homolateral and the other bilateral.

A 46. Homolateral dorsal and ventral hairs of abdominal segments one-uropod segment, not on telson or uropod. Lo- cated in area 83, it was found 9 times, and was of medium size.

A 52 responds rather tonically on stimulation of the whole of the dorsal and ventral aspects of abdominal segments 3-5 and perhaps uropod segment. Lo- cated near the stretch receptor fibers in area 78, it was found 8 times, and was of medium size.

Summary. These results give the general impression that the ventral abdominal surface is relatively unimportant as an area for touch perception. This may be related to the fact that this surface will be constantly agitated by swimmeret movements, and often by touch of the bottom. An exception is the pleural plate 1 area, which is better represented, and would also be less subject to stimulation by swimmeret movements. Even so, the contrast between the dorsal and ventral sides remains unexpectedly large. The fact that a certain part of the ventral exoskeleton was removed in order to expose the cord, cannot have played a significant role in this respect, as was shown by some con- trol experiments in which the area removed was limited to fewer segments.

Some of these fibers may play a role in the co-ordination of the metachronal rhythm of the swimmerets.

Note that fibers responding to both dorsal and ventral hairs may be identical with some of the commissure interneurons which cover large body areas, but that the evidence is insufficient to decide which ones they might be.

Interneurons integrating proprioceptive impulses (fig. 16). A single integrative interneuron has been found for the abdominal stretch receptors, which is stim-


A 24

A 35

70

43

A 2 0 I \ A 5 A 10 A 27

Fig. 16 Cross section with location of interneurons responsive to joint movements; 8, asymmetric responding interneuron.

ulated by the descending branches of the primary fibers in the 6th ganglion (Hughes and Wiersma, '60a), the others integrate swimmeret, uropod and anal valve activity.

A 61 is medium in size, fires at fre- quencies lower than the tonic stretch receptors feeding into it and is located in area 77. It was found 4 times, whereas once a similar fiber was found in the same preparation for the phasic stretch receptors.

A 20. Homolateral uropod, peripheral joints fiber. This fiber may integrate the sensory input from different joints of the uropod and can also be activated by flex- ion of the proximal part of the telson. However, such stimuli might elicit reflex contraction of a muscle receptor organ, hence the uncertainty about its nature, primary sensory or interneuron, in this and following cases in which such a mech- anism might occur. Medium in size, this fiber was found 14 times, and in most cases gave a tonic discharge. Its location is in area 84 (fig. 17).

A 68. A similar medium sized fiber, but exclusively for the basal joint of the homolateral uropod. It was found 5 times in area 77, and might well be primary (fig. 17).

A 69, also in area 77, is a partner of A 68, reacting to the same movements

of the heterolateral uropod and was noticed 6 times. It is medium in size (fig. 17).

A 70 is a large fiber which fires when either uropod is spread. Located in area 79, and found 7 times (fig. 17).

For the anal valves two special fibers have been found:

A 43. Homolateral anal valve fiber. Small, in area 82 and found 6 times, it often accompanied A 5, the asymmetric joint fiber (fig. 17). A 72 is a much larger fiber in area 76 for the heterolateral anal valve, found 5 times. Both A 43 and A 72 discharge when the valve is pulled outward, and not on mere touch (fig. 17).

As mentioned previously there are two fibers for homolateral swimmerets, which may be primary but fire only during active movement. A 35 is medium in size and is located in area 81. It fires while the 5th swimmeret is in a backward position and is inhibited by stimulation of the dorsal hairs on the same segment. It was found 8 times. A 31 similarly repre- sents the 4th swimmeret and is a neighbor of A 35 in the same area 81, is medium in size, and was obtained 16 times.

HOMOLATERAL HETEROLATERAL

Fig. 17 Ventral view indicating joints to which different interneurons respond.

224 C. A. G . WIERSMA AND G . M . HUGHES

There are three interneurons which are brought to discharge by movements of all homolateral swimmerets.

A 27 is restricted to the basal joints of the homolateral swimmerets, and in con- tradistinction to the two following fibers, it is not excited by stimulation of the anal valve. As in the two following fibers, the discharge is tonic. Located in area 83, medium in size, it was found 11 times. In males this fiber does not seem to be excited by the second swimmeret which it is in females (fig. 17).

A 10 is a medium sized fiber in area 84, which besides responding to swimmeret joints responds to the basal joint of the homolateral uropod and often the anal valve. It is sometimes difficult to distin- guish from the next fiber, when the latter does not respond to the heterolateral uropod as sometimes happens. Usually though, the difference in location is suffi- cient to decide which fiber is being recorded from. It was found 13 times (fig. 17).

A 5 is the only fiber which regularly gave an asymmetric response, reacting to all homolateral swimmerets, homolateral uropod, to the heterolateral uropod and both anal valves. It is large and very characteristically located in the medial ventral aspect of area 82. It is one of the most readily found fibers, registered 26 times and noted many other times. It appears to be identical with C 48, as was practically proven in a whole animal preparation in which it also responded to stimulation of the homolateral thoracic legs. Furthermore, this same fiber was prepared also in area 70 of the commissure in this preparation.

A 24 is a medium sized bilateral joint fiber, identical with C 46, as found in a whole animal preparation in which joint movements of all head and body appendages made this fiber fire. It was found 5 times, two of them in a whole animal. Location in area 81.

Interneurons integrating both hair and joint monement (fig. 18). The three fibers showing this type of reaction all responded to the sensory area innervated by the first root of a single abdominal ganglion.

A 7 5 Fig. 18 Cross section with location of the

interneurons with an integrative response to both hair touch and joint movement.

A 33 is a medium-sized fiber in area 81 for the homolateral third swimmeret and pleural plate 1. It was found 8 times.

A 75 is a similar fiber for the homolat- era1 5th swimmeret and pleural plate 1. It was found 5 times in area 84.

A 66 is a similar fiber for the heterolateral third swimmeret and pleural plate 1, located in area 76 and found 5 times. Similar fibers for other swimmerets are indicated, but not yet established.

Summary. The number of interneurons from joint fibers is relatively small in the cord as compared to the cornmis- sure. The approximate percentages of the total number of interneurons are 20% for the cord as against 40% for the commissure. This may be explained on the as- sumption that the swimmeret joints signal their positions to integrative areas much more by the primary sensory fibers than do other appendages. Thus the mutual co-ordination of the swimmerets, whose only really significant motion for the animal as a whole may be the metachronal beat, is largely regulated by the primary fibers. The main driving of the waves is done by interneurons (Hughes and Wiersma, 60b), so that only smaller ad- justments would have to be performed. The uropods are better represented, and i t is known that several of these interneurons reach the brain, indicating their greater importance for general co-ordination of the body.


DISCUSSION There are a number of differences be-

tween the results of this analysis of fibers in the abdominal connective and that obtained by similar methods for the circumesophageal commissure. Some of these are due to experimental circumstances but a more significant factor may be the far greater degree of local traffic in the cord than in the commissure. The many primary sensory fibers found in the cord are indicative of this difference. It is certain that in the commissure very few descending primary sensory fibers are present, whereas such fibers are abundant in the cord. The situation is less clear for ascending primary fibers since these are represented in the commissure by the many fibers from dorsal carapace hairs, whereas others which might be present, namely those from mouthparts would have been overlooked because of the experimental limitations. In the cord it is quite certain from both physiological and mor- phological evidence, that many of these fibers are thin. Thus when the locations of these bundles are superimposed on a

microphotograph of a cross section of the cord, there is a good correlation between the position of such primary sensory fiber bundles as A 9, A 18 or A 26 and the presence of fine fibers in the same areas of 84 and 82. It is strongly indicated, therefore, that all really small fibers are primary sensory ones (but not the con- verse, for the stretch receptor fibers are large as may be other primary proprioceptor fibers). If this is so, and since there are no indications that any interneuron is really small (below 10 cl), the total percentage of the larger fibers accounted for in the cord is quite considerable.

As can be seen in figure 19, out of a total of about 1200 fibers only some 370 are more than 10 p in diameter. About 68 of these now have a known function so that without considering any other fibers 20% would be accounted for, as against only 7% on the same basis in the commissure. If all entities from which single unit potentials were regularly obtained were above 20 c1 in diameter, then 50% of the population sampled by these methods will have been described physiologi-

200p - 150 100 50 0 0 50 IM)

- 150 200p

Fig. 19 Relation between number of fibers (logarithmic Y-axis) and fiber diameters in 10 p classes as counted in the left and in the right cross section of the abdomnial connectives of Procambarus clarkii on a photograph made by Dr. J. D. Robinson. Note the definite break in size between the two giant fibers and the largest ordinary fibers. Also note that the smallest fibers are more numerous than all other classes combined. Each half was counted independently.

226 C. A. G . WIERSMA AND G. M. HUGHES

cally, since there are about 140 fibers in that class. This figure is surprisingly high in relation to the other types of interneurons which must also be present. These are the descending interneurons from the thorax, the spontaneously active fibers in the cord (which are fewer than in the commissure, perhaps because all those from the higher part of the central nervous system are, of course, missing), those reacting to other sensory modalities exclusively, and finally those which though reacting, have not yet been established. It is certain that a very high pro- portion of the entities that can be found with the stimuli used has in fact been found. For in our later experiments, the number of fibers which could be desig- nated became the great majority (90% or more) of all the fibers reacting in these preparations, a situation which was not yet present in the commissure, where at best one half of such fibers could be accounted for.

If all the interneurons are indeed larger units, then any additional small fibers which may be revealed by electromicros- copy (0.2-0.7 ~1 diameter . . . J. H. McAlear, personal communication) will not affect the present issue. Larger numbers of small fibers may well be expected because of the considerable numbers of sensory fibers in the first and second roots, most of which may run either up or down or both ways for varying distances after entering the cord.

It is our view that the primary sensory fibers of the first root inflow play an im- portant part in the co-ordination of the swimmeret movements (see Hughes and Wiersma, '60b). But though this can account for the interganglionic pathways of the first root fibers, it does not explain that of the second root inflow, especially of the dorsal hair fibers. In view of the fact that for this particular area there are also many interneurons transmitting derived information, it is quite astonishing that such extensive primary fiber branch- ing should be present.

Though it is certain that many primary fibers have a restricted path length in the cord, the stretch receptor axons show that such fibers can run the full length of the central nervous system. At present, there is no direct evidence that any others do

so. But if those fibers which fire only during active backward contraction of individual swimmerets are from muscle sense organs of the type described by Alexandrowicz for the thoracic legs, finding them at distances as much as three segments from their origin may indicate that they, like the abdominal stretch receptors they resemble, also run for long distances. They would then most likely be of considerable importance in the general co-ordination of movements of the body appendages. However, if they are interneurons this reasoning would not be valid.

It is of interest to compare the number of fibers which have been found to react identically in the cord and the commissure, with the number which might have been expected from the previous work on the commissure. In this paper a total of 16 have been shown to be identical out of a possible total of 32. In addition there were about 9 fibers found which either could not definitely be shown to be identical or were found too few times to be established in the cord. Thus by far the major portion of expected fibers has been accounted for. It remains uncertain whether the other 7 are absent from the cord because of secondary integration (e.g., in the thoracic ganglia) or that further investigation will show their presence.

In the cord, as in the commissure, the great majority of interneurons integrate sensations of a single modality. Because of the larger number of elements accounted for in the cord, the conclusion that this predominance is a real feature of the crayfish nervous system, is greatly enhanced. The evidence presented here for interneurons that integrate different modalities depends on very few cases and examples of fibers which have been found relatively few times, but Kennedy and Preston ('60) also found this type of interneuron at the cord level. It is to be noted that in the commissure fibers integrating hair and proprioceptive sensation have been encountered which, as in the cord, integrate the sensations of only a single appendage.

Tract formation may be briefly discussed on a comparative basis. As in the commissure, the primary sensory fibers of


one modality form regular tracts, whether they run long distances (stretch receptors) or short ones (A 26). Furthermore, when the same type of fiber runs both forward and backward, tracts at about the same location in the cross section are present, which immediately adjoin tracts of similar sensory fibers from other segments.

Results in the cord, show that a tract may also be formed by interneurons of a certain type. In the commissure it has already been noted that the interneurons which reacted to stimulation of hairs on the homolateral thoracic appendages, run together in a tract. For axons responsive to a sensory field in a single segment, similar tracts have been noted in the abdominal connective for two sets of homolateral interneurons reacting to dorsal hairs (A 14, 15, 39, 62 and A 42, 49, 50, 56), as well as for heterolateral interneurons from these areas (A 22, 24, 30, 65).

On the other hand interneurons representing two or more segments, though similarly related among themselves, do not show this type of tract formation. Possibly this indicates that the impulses from them are not integrated with each other at a higher level, whereas those in tracts are.

The distribution of the fibers found in both commissure and cord can be compared. Their relative locations generally show a fair correspondence, but in a few cases fibers are in quite different quad- rants. Whether or not this is because a mistake has been made or that such fibers do shift in relative position, is not certain, but the latter possibility appears the more likely at present.

With regard to the nature of the sensory representation, here as in the commissure all primary sensory fibers appear to be homolateral. Homolateral interneurons are again in the great majority, but bilateral and heterolateral representation is present. Of 92 interneurons of the commissure, 63 were homolateral, 10 heterolateral, 16 bilateral and 3 asymmetric. In the cord these figures were of 57 interneurons: homolateral 39 (38), heterolat- era1 10, bilateral 7 and asymmetric 1 (2). Thus the total percentage of homolateral interneurons is about the same (68% ) in

both locations. This contradicts Prosser's belief ('35) that contralateral representation would be absent or slight in the cord.

SUMMARY 1. The electrical activity of single units

has been recorded from small bundles obtained by splitting the connectives between abdominal ganglia of the crayfish. Responses to mechanical stimulation of the abdomen, either to hairs or by movement of joints were investigated and the location of the responding units in the cross section noted.

2. An account is given of the location and properties of 75 such entities in the connective between the third and 4th abdominal ganglia, which were found a suf- ficient number of times to justify their establishment.

3. Several of these represent bundles of primary sensory fibers from a single area of the body. They may pass up or down the cord or in both directions from the ganglion which they enter. All of these remain on the same side (homolateral) but vary in the extent of their spread. This is maximal for fibers from the abdominal stretch receptors (SR1 and SR2), which send one branch to the brain and another to the last abdominal ganglion.

4. Hair fibers entering through first roots have very short intracentral pathways but those from first root propriocep- tors and second root hair fibers ascend across at least three interganglionic spaces.

5. Interneurons integrate inputs from single segments or homologous areas of several adjacent segments which may be homolater a1 , he terolater a1 , bil ater a1 , or asymmetrical in their distribution with respect to the leading off position.

6. Many have properties identical with units identified in the circumesophageal commissure but interneurons responsive to dorsal hairs on a single or two segments are a conspicuous exception, for no such fibers have been established in the commissure.

7. The great multiplicity of the central representation of a given peripheral field is one of the most striking features of this study. Its significance will only be fully understood when the efferent connections of these interneurons are investigated.

228 C . A. G. WIERSMA AND G. M. HUGHES

LITERATURE CITED Alexandrowicz, J. S. 1951 Muscle receptor or-

gans in the abdomen of Homarus vulgaris and Palinurus vulgaris. Quart. J. Micr. Sci., 92:

1958 Further observations on proprio- ceptors in Crustacea and a hypothesis about their function. J. Mar. Biol. Assoc. U. K., 37:

Allen, E. D. 1894 Studies on the nervous system of Crustacea. I. Some nerve elements of the embryonic lobster. Quart. J. Micr. Sci., 36: 461-482.

Hughes, G. M., and C. A. G. Wiersma 1960a Neuronal pathways and synaptic connexions in the abdominal cord of the crayfish. J. Exp. Biol., 37: 291-307.

1960b The co-ordination of swimmeret movements in the crayfish, Procambarus clarkii (Girard). Ibid., 37: 657-670.

163-199.

379-396.

Kennedy, D., and J. B. Preston 1960 Activity patterns of interneurons in the caudal ganglion of the crayikh. J. Gen. Physiol., 43: 655-670.

Prosser, C. L. 1935 Action potentials in the nervous system of the crayfish. 111. Central responses to proprioceptive and tactile stimulation. J. Comp. Neur., 62: 495-505.

Retzius, G. 1890 Zur Kenntniss des Nerven- systems der Crustaceen. Biol. Untersuch, N. F., I: 1-50. Central. Druck., Stockholm.

Van Harreveld, A. 1936 A physiological solution for fresh-water crustaceans. Proc. SOC. Exp. Biol. Med., 34: 428-432.

Wiersma, C. A. G. 1958 On the functional connections of single units in the central nervous system of the crayfish, Procambarus clarkii Girard. J. Comp. Neur., 110: 421471.

Wiersma, C. A. G., and R. L. C. Pilgrim 1961 Thoracic stretch receptors in crayfish and rock- lobster. Comp. Biochem. Physiol., 2: 51-64.