Fine structure of antennal sensilla of the adult mealworm beetle, Tenebrio molitor L. (Coleoptera : Tenebrionidae)

Int. J. Insect Morphol. & Embryol. 6(1): 41-60. 1977. Pergamon Press. Printed in Great Britain.

FINE STRUCTURE OF A N T E N N A L SENSILLA OF THE A D U L T MEALWORM BEETLE, TENEBRIO MOLITOR L. (COLEOPTERA " TENEBRIONIDAE)* RALPH E. HARBACH~" a n d JOSEPH R. LARSEN Department of Entomology, 320 Morrill Hall, University of Illinois, Urbana, Illinois 61801, U.S.A. (Accepted 1 November 1976)

Abstract--The types, numbers and distributions of the antennal sensilla of the adult mealworm beetle, Tenebrio molitor, were studied with the scanning and transmission electron microscopes. The types of sensilla include a sexually dimorphic bristle (sensillum chaeticum), a hair organ (sensillum trichodeum), and thin-walled, thick-walled, groow,~d, flat-tipped, and smooth-surfaced peg organs (sensilla basiconica). Hair organ~; and smooth-surfaced pegs are common on all 11 antennal segments. The other sensilla are, generally, most numerous on the distal segment, and progressively decrease in numbers on the distal inner and outer sensory fields of segments 10 through 4. Thick-., thin-walled and grooved pegs have pore systems typical of olfactory sensilla. Sensilla chaetica and thick-walled, grooved, flat-tipped, and smooth-surfaced pegs bear an apical pore suggestive of a gustatory function. Smooth-surfaced pegs, hairs, and bristles have a tubular body normally associated with mechanoreception. Index descriptors (in addition to those in the title): Receptors, sense organs, sensilla basiconica, sensillum trichodeum, sensillum chaeticum, peg organs, hair organs, bristle organs, sensory receptors, ultrastructure. INTRODUCTION THREE STUDIES o n the a n t e n n a l sensilla o f Tenebrio molitor h a v e p r e v i o u s l y a p p e a r e d in t h e l i t e r a t u r e . V a l e n t i n e (1931) o b s e r v e d 5 t y p e s o f a n t e n n a l sensilla w i t h t h e light m i c r o s c o p e a n d briefly d e s c r i b e d t h e i r e x t e r n a l s t r u c t u r e . P i e l o u (1940) r e i t e r a t e d V a l e n t i n e ' s findings a n d P i e r a n t o n i (1974) u s e d the s c a n n i n g e l e c t r o n m i c r o s c o p e to e x a m i n e the s u r f a c e t e x t u r e o f 4 o f t h e sensilla o c c u r r i n g on s e g m e n t 11. E x c e p t for P i e r a n t o n i ' s b r i e f s t r u c t u r a l e x a m i n a t i o n , t h e fi~ae s t r u c t u r e o f the sensilla has n o t b e e n i n v e s t i g a t e d p r i o r t o this study. MATERIALS AND METHODS Specimens of T. molitor were obtained from a colony maintained in our laboratory. Adult beetles 1 to 4 days of age were used in the study. Material for Lhe scanning microscope was Freon critical-point-dried (Cohen et al., 1968) in a Bomar SPC-900 critical point drying apparatus. It was subsequently mounted on aluminum stubs with Du Pont Duco cement, coated with carbon and gold-palladium on a rotary tilting stage, and examined and photographed with a Cambridge Mark II Stereoscan or a Jeolco JSM-U3 scanning electron microscope. Tissue for study in the transmission electron microscope was subjected to a 2 . 5 ~ glutaraldehyde-1 osmium tetroxic[e double fixation, buffered with 0.2 M sodium cacodylate at pH 7. The procedure, through * Part of a thesis submitted by the senior author in partial fulfillment of the requirements for the Ph.D. degree in the Department of Entomology and the Graduate College, University of Illinois, Urbana, Illinois 61801, U.S.A. 1" Present address: Department of Entomology, Box 5215, North Carolina State University, Raleigh, North Carolina 27607, U.S.A. 41

42

RALPH E. HARBACH a n d JOSEPH R, LARSEN

dehydration in 6 0 ~ ethanol, was carried out under vacuum at 4°C. The tissue was embedded in Epon 812 (Luft, 1961). Serial sections with a thickness of approximately 70 mtL were cut on a Reichert OmU2 ultramicrotome with a diamond knife, placed on formvar-coated, copper grids, and stained with 2 ~ uranyl acetate in 50 ~ ethanol. Average numbers of sensilla were determined from counts made on 108 SEM micrographs taken at a number of different magnifications. The average length and basal diameter of the external part of each sensory organ were calculated from 4 to 12 measurements, depending upon the sensillum type (Table 2), made on SEM micrographs which were taken at a minimum of 2 different magnifications. Reasonable estimates of the total number of pores on the surface of each sensillum type were obtained by multiplying: 1) the average length of each sensillum as calculated above with 2) the average number of pores occurring in 5 cross sections of known thickness taken midway along the cuticular apparatus (hair or peg) of the sensiIlum. RESULTS E a c h a n t e n n a o f T. molitor (Fig. 1) has a n a v e r a g e l e n g t h o f 3.5 m m a n d c o n s i s t s o f 1 ! s e g m e n t s . E a c h s e g m e n t is s w o l l e n d i s t a l l y ; a d o r s o v e n t r a l f l a t t e n i n g o f the distal s e g m e n t s c a u s e s g r e a t e r swelling laterally. T h e lateral e x p a n s i o n s are r e s p o n s i b l e for the slowly i n c r e a s i n g t r u n c a t i o n o f s e g m e n t s 5 t h r o u g h l0 a n d the serrate a p p e a r a n c e o f the a n t e n n a . S e g m e n t 11 is r o u n d e d a n d b r o a d at the t o p a n d a t t e n u a t e at the base. Sensilla a r e c o m m o n o n all o f the a n t e n n a l s e g m e n t s (Fig. 1). T h e y are m o s t h i g h l y c o n c e n t r a t e d o n s e g m e n t 11 (Fig. 3), t h e s w o l l e n distal i n n e r s u r f a c e o f s e g m e n t s 5 t h r o u g h 10 (Figs. 4-9), a n d the swollen distal o u t e r s u r f a c e o f s e g m e n t s 8 t h r o u g h l0 (Figs. 4-6). T h e size o f t h e s e a r e a s o r fields, a n d the n u m b e r o f sensilla l o c a t e d o n t h e m , s t e a d i l y d e c r e a s e s as o n e m o v e s p r o x i m a l l y f r o m s e g m e n t 11 to s e g m e n t 5 ( T a b l e 1; Figs. 3-9). A distal o u t e r s e n s o r y field is n o t p r e s e n t o n s e g m e n t 7 o r the m o r e p r o x i m a l s e g m e n t s (Fig. 1). A h i g h l y r e d u c e d distal i n n e r s e n s o r y field o c c u r s o n s e g m e n t 4 (Fig. 10), b u t n o t o n the m o r e p r o x i m a l s e g m e n t s (Fig. 1). TABLE I. AVERAGE NUMBERS OF SENSILLA ON AN ANTENNA OF ADULT TENEBRIO MOLITOR

Antennal segment 11 10 9 8 7 6 5 4 3 2 1

ThickThinFlatwalled walled Grooved tipped peg organs peg organs peg organs peg organs 196 62 46 23 ----. . .

1338 705 455 208 118 34 3 -. . .

30 16 ll 7 5 4 2 1 . . .

. . .

16 8 5 2 1 2 1 2

Smoothsurfaced peg organs 6 7 7 14 18 17 20 20 26 8 15

S. trichodea 148 83 73 81 89 87 81 82 115 48 102

S. chaetica 23 13 13 12 9 9 5 1~(3~*) ----

* These have the size and appearance of smooth-surfaced pegs. They are located, however, in a curved row as are S. chaetica on the sensory fields of other segments. E a c h a n t e n n a b e a r s 7 d i s t i n c t s e n s i l l u m types. A c c o r d i n g to C a l l a h a n ' s (1975) classification, t h e s e n s i i l u m t y p e s are as f o l l o w s : A. B.

S e n s i l l u m c h a e t i c u m (bristle o r g a n ) (Figs. l 1, 20, 2 3 - 2 6 ) S e n s i l l u m t r i c h o d e u m (hair o r g a n ) (Figs. 15, 27-30)

Fine Structure of Antennal SensiUa of the Adult Mealworm Beetle

PLATE 1. FIG. 1. Dorsal surface of right antenna of female. Compare bristles at arrows with male spatulate bristles at arrows in Fig. 2. FIG. 2. Dorsal surface of segments 5-9 of right antenna of male.

43

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RALPH E. HARBACHand JOSEPH R. LARSEN

PLATE 2. FtGS. 3-6. Distal view of antennal segments 11-8, respectively. Observe decrease in number of sensilla proximally from segment I 1 to segment 8. Thick-walled pegs are common on distal outer sensory field while there are few on distal inner field. FIG. 3. Segment 11 of right antenna of male. FiG. 4. Segment 10 of right antenna of female. FIG. 5. Segment 9 of right antenna of male. F~6. 6. Segment 8 of left antenna of male. Note that spatulate bristles are located on ventral side of inner field only. Compare with right antennal segments of Figs. 5, 7, 8.

6.3/zm

4.0/zm

3.0 #m

4.1/zm

5.8 tLm 15.5/~m

25.0/~m 50.6 t~m

Thick-walled peg organ

Thin-walled peg organ

Grooved peg organ

Flat-tipped peg organ

Smooth-surfaced peg organ S. trichodeum

S. chaeticum Male spatulate bristle

range

19.4-31.3 tzml 37.8-60.0 t~m

5.5-6.0 t~m 8.7-31.9/~m ~

4.0-4.3/zm

2.6-3.3 t,m

2.7-7.1/zm

4.4-9.8/zm

Length

4.6 ~m 4.6 t~m

1.7 t~m 1.7 t~m

1.6 t~m

1.6 tma

2.0 t~m

1.6/~m

4.3-5.2 t~m 3.9-6.3 t~m

1.6-2.0 t~m 0.8-3.2/zm

1.4-1.8 t~m

1.2-1.8 t,m

1.6-2.3 t~m

1.3-1.9/zm

Basal diameter mean range

12 10

5 12

4

6

11

10

blunt with pore flattened with pore

blunt with pore pointed

blunt with pore

blunt with pore

blunt

blunt with pore

Tip

TENEBRIO MOLITOR

No. measured

E X T E R N A L CHARACTERISTICS OF ANTENNAL SENSILLA OF ADULT

* Progressively decrease in length distally along antenna. "~Progressively decrease in length proximally along antenna.

mean

Type

TABLE 2.

smooth or slightly corrugated with pores smooth or slightly corrugated with pores strongly corrugated with pores smooth and corrugated smooth smooth or helically corrugated corrugated corrugated

Surface

~"

;~

=z

,~ = =

= ?,

~"

46

RALPH E. HARBACHand JOSEPH R. LARSEN C. 1. 2. 3. 4. 5.

Sensilla b a s i c o n i c a (peg o r g a n s ) S m o o t h - s u r f a c e d p e g (Figs. 18, 19, 31,32) T h i c k - w a l l e d p e g (Figs. 14, 33, 35) T h i n - w a i l e d p e g (Figs. 13, 34, 35) F l a t - t i p p e d p e g (Figs. 16, 36-38) G r o o v e d peg (Figs. 17, 39-44) TABLE 3.

INTERNAL

Type

C H A R A C T E R I S T I C S OF A N T E N N A L S E N S 1 L L A O F A D U L T

Ave. no. pores

Walls

Thick-walled peg organs Thin-walled peg organ Grooved peg organ

thick with pores thin with pores thick with pores with grooves

TENEBRIO ,VIOLITOR

lnnervations

Dendritic sheath

2

thin

450 (1 at tip)

Probable function gustatory; olfactory

1089 214 (1 at tip)

2

thin

5

thick thick intermediate thickness intermediate thickness intermediate thickness

Flat-tipped peg organ Smooth-surfaced peg organ

thick

(1 at tip)

2

thick

(1 at tip)

6

S. trichodeum

thick

--

1

S. chaeticum

thick

(1 at tip)

6

Male spatulate bristle

thick

(1 at tip)

6

olfactory gustatory; olfactory; hygroreceptor gustatory tactile; gustatory

tactile tactile; gustatory tactile; intermediate gustatory; thickness pheromone receptor

A m o d i f i e d s e n s i l l u m c h a e t i c u m ( s p a t u l a t e bristle) o c c u r s o n the m a l e a n t e n n a e (Figs. 2, 12, 21-26). T h e e x t e r n a l a n d i n t e r n a l c h a r a c t e r i s t i c s o f e a c h s e n s i l l u m t y p e are s u m m a r i z e d in T a b l e s 2 a n d 3 a n d i l l u s t r a t e d by the figures d e s i g n a t e d a b o v e . F o r c o m p a r i s o n , t h e n a m e s u s e d in this a n d t h e studies o f V a l e n t i n e (1931), P i e l o u (1940), a n d P i e r a n t o n i (1974) to d e n o t e the s a m e sensilla a r e listed in T a b l e 4. TABLE 4.

T H E D I F F E R E N T N A M E S U S E D I N T H E P R E S E N T A N D E A R L I E R S T U D I E S T O D E N O T E T H E SAME S E N S I L L U M TYPES WHICH

Present Study

OCCUR ON THE ANTENNAE OF

Valentine (1931)

sensillum chaeticum sensillum trichodeum thick-walled peg organ thin-walled peg organ grooved peg organ flat-tipped peg organ smooth-surfaced peg organ -

-

bristle hair organ * peg organ --pit peg organ enclosed peg organ

* Probably classified as a peg organ.

TENF.BRIO MOLITOR

Pielou (1940) bristle or spine hair sensillum * peg organ (basiconic sensillum) --pit peg organ (coeloconic sensillum) enclosed peg organ (ampullaceous sensillum)

Pierantoni (1974) D receptor C receptor B receptor A receptor M

Fine Structure of Antennal Sensilla of the Adult Mealworm Beetle

PLATE 3. FIGS. 7-10 Inner surface of antennal segments 7-4, respectively. Observe proximally decreasing number of sensilla on distal inner field. A distal outer field is absent from these segments and more proximal ones. Distal inner field of segment 4 is highly reduced. It usually has only a few peg organs (flat-tipped and grooved) and, in males, 1 or 2 sensilla chaetica. A distal inner field is absent from more proximal segments (see Fig. 1). FIG. 7. Segment 7 or right antenna of male. FIG. 8. Segment 6 of right antenna of male. FIG. 9. Segment 5 of right antenna of female. FIG. 10. Segment 4 of left antenna of male.

47

48

RALPH E. HARBACHand JOSEPH R. LARSEN

PLATE 4.

Fine Structure of Antennal Sensilla of the Adult Mealworm Beetle

49

Sensillum chaeticum (bristle organ) Sensilla chaetica are confined to segment 11, the distal inner sensory fields of segments 4 through 10, and the distal outer fields of segments 8 through 10. Progressing proximally from segment 11, each succeeding segment bears fewer of these sensilla (Table 1; Figs. 1, 3-10). Each of the sensory fields on segments 5 through 10 has a row of sensilla chaetica. The sensilla are more or tess evenly spaced along the row. The row curves around the periphery of each sensory field (Figs. 4-9). The primary location of the male spatulate bristles is on the ventral portion of the distal inner fields of segments 4, or 5 through 9, or 10 (Figs. 2, 5-8). Progressing from segment 11 to the more proximal segments, normal bristles decrease in length. The decrease is more pronounced on female antennae (Fig. 1), since male antennae have fewer of these bristles. In the female, the smallest of these receptors, especially those on segment 4, resemble the smooth-surfaced peg (see below) in both size and appearance. The male spatulate bristle and the other bristle differ only in length and the form of their tip. The normal bristle has a mean length of 25/~m and the male bristle has a mean length of 50.6/xm. The tip of the normal bristle is blunt (Fig. 11); that of the male structure is typically flattened and expanded (Fig. 12). Both are straight or slightly curved structures with a corrugated surface (Figs. 11, 12), an apical pore (Fig. 22), and a very thick wall (Figs. 20, 21). Both have a well-defined socket (Fig. 11). Six dendrites innervate the sensillum chaeticum (Figs. 24-26). A single dendrite forms a tubular body (Fig. 24) which inserts into the cuticle at the base of the bristle. The remaining dendrites penetrate the lumen of the bristle. At least one of these traverses the length of the lumen and terminates in the region of the apical pore. The others terminate at various intervals along the length of the lumen (Fig. 20). The dendrites normally remain unbranched, but those of the male spatulate bristle occasionally bifurcate in the region of the expanded tip. The dendrites or dendritic branches are separated from the external environment at the site of the apical pore by a pore plate (Fig. 22). Above the ciliary region, the dendrites are encased within a dendritic sheath of intermediate thickness (Figs. 24-26). As the dendrites approach the base of the bristle, they begin to be separated by invaginations of the sheath (Fig. 26). The invaginations progressively extend more deeply inward (Fig. 25) until, at a point immediately below the bristle, they completely or almost completely partition the dendrites (Fig. 24). The dendritic sheath apparently loses its invaginations and becomes thinner as it enters the lumen of the

FIG. 11. Bristle located near apex of segment ll. Note its corrugated surface, deep socket, and pores of dermal glands located around its base. FIG. 12. Spatulate bristle of male. These receptors are common on segments 5-9. Note their flattened and expanded apex and their corrugated surface. FIG. 13. Thin-walled peg. Note its finely corrugated surface. FIG. 14. Thick-walled peg. It has a corrugated surface and a tapered but blunt tip which bears an apical pore (not visible). FIG. 15. Hair organ. This sensillum type has a corrugated surface, but it cannot be discerned in this scanning electron micrograph. FIG. 16. Flat-tipped peg. Note its slender and tapering form and moderately deep corrugation. Its tip has a flattened, puckered appearance and bears an apical pore. Fm. 17. Grooved peg. It has an apical pore and a deeply corrugated or grooved surface. FIG. 18. Smooth-surfaced peg. Note that peg has a smooth surface, apical pore, and deep socket. It is always slightly curved and non-tapered. FIG. 19. Higher magnification of tip of smooth-surfaced peg showing apical pore. lMAE 6 / | - - D

50

RALPH E. HARBACH a n d JOSEPH R. LARSEN

PLATE 5.

Fine Structure of Antennal Sensilla of the Adult Mealworm Beetle

51

bristle. It separates the lumen into 2 chambers (Fig. 23) before it completely fuses with the wall of the bristle a short distance above the socket of the sensillum.

Sensillum trichodeum (hair organ) Hairs progressively decrease in length distally along the antenna, but they are evenly dispersed over the surface o f each antennal segment (Fig. 1). Segments 1, 3, and 11, which have a greater surface area than the other segments, possess the greatest numbers of sensilla trichodea (Table 1). Segment 2 bears the smallest n u m b e r of these organs, but it also has a smaller surface area than the other segments (Table 1 ; Fig. 1). Segments 4 t h r o u g h 10 are o f nearly equal size (Fig. 1). The mean n u m b e r of sensilla trichodea on each o f these segments is 82 (Table 1). The hair is a long, slender, pointed structure with a s m o o t h or helically corrugated surface (Fig. 15). It has a deep socket (Fig. 27) which is c o m m o n l y surrounded by the pores of dermal gland,; (Fig. 15). A single dendrite innervates the sensillum (Figs. 28-30), but it does not enter the small lumen of the thick-walled hair (Fig. 27). Instead, it forms a tubular b o d y which inserts into the cuticle at the base of the hair (Fig. 28). A dendritic sheath of intermediate thickness (Figs. 28-30) encases the outer segment o f the dendrite ;above the ciliary region. Between the point where the dendrite enters the sheath and the region of the tubular body, the sheath invaginates into the dendrite (Figs. 29, 30). The invaginations are most extensive just below the tubular b o d y (Fig. 29).

Sensillum basi~conicum (smooth-surfaced peg organ) Smooth-surfaced pegs, although they occur on all 11 segments, are not very numerous. They occur in lower numbers on the most distal segments (Table 1). Fewer o f these pegs occur on the inner surfaces of segments 4 t h r o u g h 11 than on their outer surfaces, but they are evenly disi~ersed on segments 1 t h r o u g h 3 (Fig. 1). The pegs are absent from the proximally attenuated part of each of the 11 segments. The peg is slightly curved but not tapered (Fig. 18). Its tip is blunt and bears an apical pore (Figs. 18, 19). The peg rests in a b r o a d and deep depression (Figs. 6-10) and has a well-defined socket (Fig. 32). This peg, like the sensillum chaeticum, is innervated by 6 dendrites. The outer segment of one of the dendrites has a tubular b o d y which inserts into the cuticle at Lhe base o f the peg (Fig. 31). The remaining 5 dendrites enter the lumen of the

FIGS. 20-26. Sensillum chaeticum. FIG. 20. Cross section midway along length of bristle. At this point, only 3 dendrites are found in lumen. FIG. 21. C:;oss section through flattened tip of male spatulate bristle. Dendrites are contained within a lumen which is located at one side of flattened tip. F~. 22. Section through pore area of spatulate bristle. Pore is located at side of flattened tip closest to dendrites. A pore plate separates pore canal from dendrites. F~6. 23. Higher magnification of lumen of bristle. At this point, dendritic sheath divides lumen into 2 chambers and 5 dendrites are present (cf. Figs. 20, 21). FIG. 24. Section immediately below base of bristle showing 6 dendrites before they innervate lumen. Note that dendritic sheath partitions dendrites. One dendrite forms a tubular body which inserts into cuticle at base of bristle. FIG. 25. Section below region of tubular body. Large dendrite gives rise to tubular body. Note that dendrites are still partitioned by dendritic sheath at this level. Flo. 26. Section through epidermal cell complex showing normal enfolded relationship of trichogen and tormogen cells. At this level, dendrites are indistinguishable and dendritic sheath is only beginning to separate dendrites.

52

RALPH E. HARBACHand JOSEPH R. LARSEN

PLATE 6.

Fine Structure of Antennal Sensilla of the Adult Mealworm Beetle

53

peg (Fig. 32), but it is uncertain whether or not they all extend to the tip o f the peg. The distal processes o f the dendrites are enclosed within a dendritic sheath o f intermediate thickness (Fig:. 31). It is uncertain, but the dendrites m a y be completely partitioned by the sheath along its entire length. The sheath fuses with the wall of the peg in the region o f the socket (Fig. 32).

Sensillum basiconicnm ( thick-walled peg organ) Thick-walled pegs occur primarily on segments 8 t h r o u g h 11. They are located chiefly on the outer surface o f these segments, but a few can also be found on their inner surface. The sensilla are most numerous on segment 11. They progressively decrease in n u m b e r and are least n u m e r o u s on segment 8 (Table 1 ; Figs. 3-6). A few of these receptors are occasionally f o u n d on the distal inner fields o f segments 5 t h r o u g h 7. This peg organ has a mean length o f 6.3/zm. It has a poorly defined socket, a s m o o t h or corrugated surface, and a blunt tip which bears an apical pore (Fig. 14). The peg has a thick wall fro~x~ which it gets its name. The wall is perforated by approximately 450 pores, and each lead.s into a canal which contains a system of tubules. The pore-tubules m a k e contact with dendritic branches within the lumen o f the peg (Fig. 33). The sensillum is innervated by 2 dendrites (Fig. 35). These bifurcate several times as they enter the lumen of the peg to give rise to a n u m b e r o f dendritic branches. Only a few of the branches, however, run the entire length o f the lumen to terminate in the vicinity o f the apical pore. Above the ciliary region, the dendrites enter a thin dendritic sheath (Fig. 35). The sheath erLds at the base of the peg, but its exact relationship to the peg has not been observed. The sheath varies in form along its length. It often has an irregular appearance with outer folds or protuberances (Fig. 35). It may partially or completely partition the dendrites in different regions.

Sensillum basiconicum ( thin-walled peg organ) Thin-walled pegs are the most numerous of the sensory organs. They are evenly spaced but crowded on segment 11, the distal inner fields o f segments 5 t h r o u g h 10, and the distal outer fields of' segments 8 t h r o u g h 10 (Figs. 3-9). They are most n u m e r o u s on segment 11. Proceeding proximally from the eleventh segment, each succeeding segment, t h r o u g h 5, bears approximately one-half as m a n y o f these organs as the segment immediately distal to it (Table 1).

F~Gs. 27-30. Sensillum trichodeum. FIG. 27. Section through very base of hair showing its socket. Note that lumen of hair is not innervated. FIG. 28. Section immediately below base of hair. Tubular body of single dendrite can be seen as it begins to be embedded in cuticle at base of hair. FIG. 29. Seztion below region of tubular body. At this point, dendrite is invaginated by dendritic sheath. FIG. 30. S~ztion through region where dendrite is beginning to be invaginated by dendritic sheath. Observe large number of mitochondria in trichogen cell. FIGS. 31, 32. Smooth-surfaced peg organ. Micrographs are from segment 3. FIG. 31. Longitudinal section through dendrites and dendritic sheath. Six dendrites innervate peg. One has tubular body which inserts into cuticle at base of peg. FIG. 32. Longitudinal section through peg. Observe deep socket, dendrites as they emerge from dendritic sheath, and absence of pores in wall of peg.

54

RALPH E. I-IARBACHand JOSEPH R. LARSEN

PLATE 7.

Fine Structure of Antennal Sensilla of the Adult Mealworm Beetle

55

The external, cuticular part of this sensillum has a m e a n length o f 4/xm. It rests in a depression which is deeper and has steeper walls than those which surround the other sensillum types that occur on the distal outer and inner fields. Its surface is s m o o t h or slightly corrugated (Fig. 13) and bears numerous pores. Its tip is r o u n d e d or domed, and lacks an apical pore. The lumen of the peg is innervated by 2 dendrites (Fig. 35). These bifurcate repeatedly in the basal region of the peg to give rise to m a n y dendritic branches (Fig. 34). The branches terminate at various points along the length of the lumen with successively fewer reaching the apex of tlze peg. The thin wall of the peg (Fig. 34) is perforated by approximately 1089 pores. Each pore opens into about 7 tubules which enter the lumen to make contact with the membranes o:F"the dendritic branches. The 2 inne:rvating dendrites are encased within a thin dendritic sheath (Fig. 35) which proceeds from a point just above the ciliary regions of the dendrites to the base o f the peg. As is the case with the thick-walled peg, the sheath is irregular in shape, m a y occasionally separate the d.endrites, and has an u n k n o w n relationship with the base of the peg.

Sensillum basiconicum (flat-tipped peg organ) Fewer than 40 flat-tipped pegs occur on each antenna. Almost one-half o f these are f o u n d along the apical margin of segment 11 (Fig. 3); the rest are primarily f o u n d on the distal inner fields of segments 10 t h r o u g h 4 (Table 1). A few also occur on the distal outer field o f segment 10. At least one but as m a n y as 4 m a y occur on segment 4 in females. At least one is usually present on segment 4 in males (Fig. 10). The peg has a m e a n length o f 4.1 /xm. Its surface is usually corrugated (Figs. 16, 36), but its base and distal one-fourth to one-third may be s m o o t h surfaced (Fig. 16). The tip o f the peg is flat, usually appears puckered, and bears an apical pore (Fig. 16). The wall is thick and does not contain pores (Fig. 36). Two dendrites innervate the peg and completely fill its lumen (Figs. 36-38). It is believed that they both terminate at the site o f the apical pore. The dendrites are encased within a thick dendritic sheath which increases in thickness as it approaches the base o f the peg (Fig. 38). The. sheath fuses with and becomes part of the wall of the peg (Fig. 37). There often appears to be m o r e than 2 dendrites inside the dendritic sheath (Figs. 37, 38). This p h e n o m e n o n is believed to be caused by invaginations o f the sheath (Fig. 37)

FIG. 33. Cross section through thick-walled peg. Only a few dendritic branchesextendtotip of peg. Observe pore-tubule systems. FiG. 34. Slightly oblique section through peg of thin-walled peg. Observe thin wall with poretubule syst,ems. There are approximately 7 tubules per pore which make contact with dendritic membranes. Contacts cannot be seen in this micrograph. FIG. 35. Section through epidermal cell complex of thin and/or thick-walled pegs. Two dendrites which inne:wate pegs are encased within very thin dendritic sheath. Sheath often has a number of folds or protuberances. FtGs. 36-38. Flat-tipped peg organ. FIG. 36. Cross section midway along peg. Two dendrites are always found in lumen. Peg lacks pores and/or channels of any sort. Note surface corrugations. FIG. 37. Section just below base of peg showing portion of socket. Note that an extremely thick dendritic sheath has fused with wall of peg. More than 2 dendrites appear to be present due to invaginations of dendritic sheath. FIG. 38. Section below peg. Note thick dendritic sheath with invaginations. Sheath becomes thinner below this region.

56

RALPH E. HARBACHand JOSEPH R. LARSEN

PLATE 8. FIGS. 39-44. Grooved peg organ. FIGS. 39, 40. Cross sections through peg. Peg is innervated by 5 dendrites (Figs. 41, 43, 44), but only one usually reaches distal end of peg (Fig. 39). Occasionally a dendrite may bifurcate within lumen (Fig. 40). Observe deep grooves in surface. These communicate with radial channels which appear to contain a dense strand or dendritic protrusion (Fig. 40). FIG. 41. Five dendrites encased within thick dendritic sheath just below region where it fuses with peg wall. FiG. 42. Higher magnification of a cross section of peg. Observe the radial channel which contains a dense strand and has a dendritic protrusion entering it. FIGS. 43, 44. Sections progressing to deeper levels below peg. Note that dendritic sheath becomes thinner as it approaches ciliary region of dendrites (cf. Fig. 41).

Fine Structure of Antennal Sensilla of the Adult Mealworm Beetle

57

between the dendrites and their branchlike extensions. No more than 2 dendrites have been observed below the dendritic sheath. Sensillum basieonieum (grooved peg organ) Grooved pegs occur only on the distal 8 segments of the antenna. Nearly one-half of the total number of these organs are located on segment 11 ; the rest are found in decreasing numbers on ~,egments 10 through 4 (Table 1). The latter are primarily confined to the distal inner fields of the segments, but a few also occur on the distal outer fields of segments 9 and 10. None, one, or 2 may occur on segment 4. The grooved peg is the smallest of the antennal receptors. The peg has a mean length of 3/xm. It is innervated by 5 dendrites (Figs. 41, 43, 44) which are tightly clustered and nearly fill its lumen. They generally remain unbranched, but an occasional bifurcation is observed (Fig. 10). Usually only one dendrite reaches the region of the apical pore (Fig. 39). The other dendrites terminate at successively lower levels within the tapered lumen. The wall of the peg is perforated by approximately 214 pores. These are unlike those of typical olfactory organs because they do not open into tubule systems. Instead, they open into a radial channel which often contains a dense strand (Fig. 40). The strands approach the dendrites, but their exact relationship to the dendritic membranes is unknown. There is evidence, however, that dendritic protrusions enter the radial channels (Fig. 42). The protrusions n'Lay be unrelated to, contiguous with, or in contact with the strands. The dendrkes enter a dendritic sheath just above their ciliary regions (Fig. 44). At this point, the sheath is thin, but it steadily increases in thickness until it reaches the base of the peg (Figs. 41, 43, 44). It fuses to the wall of the peg a short distance above the poorly defined socket of the sensillum. Enclosed peg crgan Valentine (1931) and Pielou (1940) reported that enclosed peg organs occurred on all segments of the antenna, but their presence was not confirmed in the present SEM and TEM investigation. The various types of pegs, however, appear to be surrounded by cuticle when they are sectioned at certain angles. This apparently occurs when a section is taken through the wall of the depression which surrounds the base of each peg. it is believed that this phenomenon led Valentine and Pielou to infer the presence of enclosed pegs from similar results obtained with preparations for the light microscope. DISCUSSION The present study shows that there are 7 morphologically distinct sensilla on the antennae of T. molitor: a sexually dimorphic bristle organ, a hair organ, and thick-walled, thinwalled, smooth-surfaced, grooved, and fiat-tipped peg organs. The first 5 of these types were previou..dy known by different names from the studies of Valentine (1931), Pielou (1940), and Pierantoni (1974). Since the grooved and fiat-tipped pegs cannot be distinguished from the thin-walled pegs in the light microscope, Valentine and Pielou were probably unable to observe them. Based on purely morphological evidence, it is likely that the thin- and thick-walled peg organs have an olfactory function. These sensilla are perforated by numerous pores which contain a nuraber of tubules making direct contact with dendritic membranes. Since it was first hypothesized that pore tubules serve to conduct olfactory molecules to the trans-

58

RALPH E. HARBACH a n d JOSEPH R. LARSEN

duction sites on the receptor membranes (Steinbrecht and Mfiller, 1971), tubule-membrane contacts have been observed in the olfactory organs of a number of insects (Borg and Norris, 1971 ; Norris and Chu, 1974; Harbach and Larsen, 1976). The grooved peg organ may function as an olfactory receptor as well. Its pores, however, do not open into a series of tubules. Instead, each pore opens into a radial channel which contains a dense strand. Although grooved pegs with a similar morphology have been studied in Sareophaga argyrostoma (Slifer and Sekhon, 1964), Locusta migratoria (Steinbrecht, 1969), Cimex lectularius (Levinson et al., 1974), Aedes aegypti (McIver, 1974), and Oncopeltusfasciatu~ (Harbach and Larsen, 1976), the dense strands have never been shown to have a tubular structure like the tubules of typical olfactory sensilla. In spite of this, grooved pegs are known to function as olfactory receptors (Lacher, 1967; Steinbrecht, 1969; Kellogg, 1970). In Locusta migratoria, grooved pegs have been shown to have a hygroreceptive function (Waldow, 1970) in addition to an olfactory function (Boeckh, 1967; Steinbrecht, 1969). Altner et al. (1973) also showed that a peg of similar morphology on the antennae of Periplaneta americana possesses hygroreceptive units. In addition, the humidity responses of Oncopeltusfasciatus closely parallel the distribution of antennal, grooved pegs (Harbach, unpublished observations) which are similar in structure (Harbach and Larsen, 1976) to those in Tenebrio. On the basis of these findings, it is possible that the grooved peg of Tenebrio has a hygroreceptive function. Olfactory organs on the antennae of Oncopeltusfasciatus (Harbach and Larsen, 1976) and a number of fulgoroid insects (Marshall, 1973) have dendritic extensions which enter the pore canals. The results of the present study indicate that similar protrusions may be unrelated to, contiguous with, or in contact with the dense strands. If they are in contact, the strands may be functionally synonymous with the pore tubules of other olfactory sensilla. The sensilla chaetica and the thick-walled, grooved, flat-tipped, and smooth-surfaced pegs probably function as gustatory organs. Each of these sensilla has an apical pore. Although there is no evidence from the present morphological investigation to indicate that the apical pores of these sensilla are capable of being opened and closed, an openingclosing process is known for the terminal pores of gustatory sensilla in Schistocerca gregaria (Blaney and Chapman, 1969), Locusta migratoria (Blaney et al., 1971), and the blow flies Phormia regina and Calliphora L~icina (Stiirckow et al., 1973). Although the location of the flat-tipped, smooth-surfaced, thick-walled, and grooved pegs beneath the longer bristles and hairs may or may not prevent them from coming into contact with the grain or grain products in which the beetle lives, the fact that the dendrites of these pegs communicate with the external environment via an apical pore indicates that they could be responsive to air-borne stimuli. Recently, St~dler and Hanson (1975) demonstrated that the gustatory receptors of larval Manduca sexta respond to the odors of natural food substances. This is not mentioned here to indicate that the probable gustatory receptors of Tenebrio function as "olfactory" organs, but in order to show that the receptors could react to molecules transported in air. The male spatulate bristle may have a specialized gustatory function. The primary location of these organs on the ventral portion of the distal inner fields of segments 4, or 5 through 9, or 10 indicates that they may function as contact pheromone receptors. Insect mechanoreceptors normally possess a dendrite which has a distal tubular body (McIver, 1975). Since a tubular body is present in the sensillum chaeticum, sensillum

Fine Structure of Antennal Sensilla of the Adult Mealworm Beetle

59

t r i c h o d e u m , a n d s m o o t h - s u r f a c e d p e g o r g a n , t h e s e r e c e p t o r s a r e likely t o h a v e a m e c h a n o receptive function. Acknowledgments--We extend our thanks to Mrs. Mary Fisher for her technical assistance and for proofreading the manttscript. This study was supported by a USPH Training Grant, G M 1076.

REFERENCES ALTNER, H., K. S. ERNST, I. KOLNBERGER and R. LOETUS. 1973. Feinstruktur und ad/iquater Reiz bei Insektensensillen mit Wandporen. Verb. Dtsch. Zool. Ges. 66: 48-53, BLANEY,W. M. and R. F. CHAPMAN. 1969. The fine structure of the terminal sensilla on the maxillary palps of Schi~tocerca gregaria (Forskfil) (Orthoptera, Acrididae). Z. Zellforsch. Mikrosk. Anat. 99: 74-97. BLANEY, W. M., R. F. CHAPMAN and A. G. CooK. 1971. The structure of the terminal sensiUa on the maxillary palps of Locusta migratoria (L.), and changes associated with moulting. Z. Zellforsch Mikrosk. Anat. 121: 48-68. BOECKH, J. 1967. Reaktionsschwelle, Arbeitsbereich und Spezifit/it eines Geruchsrezeptors auf der Heuschreckenantennae. Z. Vgl. Physiol. 55: 378-406. BORG, T. K. and D. M. NORRIS. 1971. Ultrastructure of sensory receptors on the antennae of Scolytus multistriatus (Marsh.). Z. Zellforsch. Mikrosk. Anat. 113: 13-28. CALLAHAN,P. S. 1975. Insect antennae with special reference to the mechanism of scent detection and the evolution of the sensilla. Int. J. Insect Morphol. EmbryoL 4: 381-430. COHEN, A. L., D. P. MARLOW and G. E. GARNER. 1968. A rapid critical point method using fluorocarbons ("Freons") as intermediate and transitional fluids. J. Microsc. (Paris) 7: 331-42. HARBACH, R. E. and J. R. LARSEN. 1976. Ultrastructure of sensilla on the distal antennal segment of adult Oncope!tus fasciatus (Dallas) (Hemiptera: Lygaeidae). Int. J. Insect Morphol. Embryol. 5: 23-33. KELLOGG,F. E. 1970. Water vapour and carbon dioxide receptors in Aedes aegypti. J. Insect Physiol. 16: 99-108. LACHER, V. 1967. Elektrophysiologische Untersuchungen an einzelnen Geruchsrezeptoren auf den Antennen weiblicher Moskitos (Aedes aegypti L.). J. Insect Physiol. 13: 1461-70. LEVINSON, H. Z., A. R. LEVINSON, B. M/3LLER and R. A. STEINBRECHT. 1974. Structure of sensiUa, olfactory perception, and behaviour of the bedbug, Cimex lectularius, in response to its alarm pheromone. J. Insect Physiol. 20: 1231-48. LUFT, J. 1961. Improvements in epoxy resin embedding methods. J. Biophys. Biochem. Cytol. 9: 409. MARSHALL,A. T. 1973. Vesicular structures in the dendrites of an insect olfactory receptor. Tissue Cell 5: 233-41. MCIVER, S. B. 1974. Fine structure of antennal grooved pegs of the mosquito, Aedes aegypti. Cell Tissue Res. 153: 327-37. MCIvER, S. B. 1975. Structure of cuticular mechanoreceptors of arthropods. Annu. Rev. Entomol. 20: 381-97. NORRIS, D. M. and H. CHU. 1974. Morphology and ultrastructure of the antenna of male Periplaneta americana as related to chemoreception. Cell Tissue Res. 150: 1-9. PIELOU, D. P. 1940. The humidity behaviour of the mealworm beetle, Tenebrio molitor L. J. Exp. Biol. 17: 286-94. PIERANTONI, R. '~974. Electron scanning microscopy of the antennal receptors in Tenebrio molitor. A steroscopic analysis. Cell Tissue Res. 148: 127-42. SLIFER, E. H. a r d S. S. SEKHON. 1964. Fine structure of the sense organs on the antennal flagellum of a flesh fly, Sarcophaga argyrostoma R.-D. (Diptera, Sarcophagidae). J. Morphol. 114: 185-208. SrXDLER, E. and F. E. HANSON. 1975. Olfactory capabilities of the "'gustatory" chemoreceptors of the tobacco hornworm larvae. J. Comp. Physiol. 104: 97-102. STEINBRECHT,R. A. 1969. Coroi-arative morphology of olfactory receptors, pp. 3-21. In C. PFAFFMANN(ed.) Olfaction and Taste. Rockefeller University Press, New York. STEINBRECHT, R. A. and B. MULLER. 1971. On the stimulus conducting structures in insect olfactory receptors. Z. ZellJbrsch. Mikrosk. Anat. 117: 570-5. ST/.JRCKOW, B., P. E. HOLBERT, J. R. ADAMSand R. J. ANSTEAD. 1973. Fine structure of the tip of the labellar taste hair of the blow flies, Phormia regina (Mg.) and Calliphora vicina R.-D. (Diptera, Calliphorid~Le). Z. Morphol. Tiere 75: 97-109. VALENTINE,J. M. 1931. The olfactory sense of the adult mealworm beetle Tenebrio molitor (Linn.). J. Exp. Zool. 58: 165-227. WALDOW, U. 1970. Elektrophysiologische Untersuchungen an Feuchte-, Trocken- und Kfi.lterezeptoren auf der Antennae der Wanderheuschrecke Locusta. Z. l/gl. Physiol. 69: 249-83.

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ap B1 B2 B4 B5 C co cu d dg do dp ds f g in mt ot pc pp ptu rc s SC sf st T tb tc tcv to ve

RALPH E. HARBACH and JOSEPH R. LARSEN ABBREVIATIONS USED IN FIGURES apical pore thick-walled sensillum basiconicum thin-walled sensillum basiconicum flat-tipped sensillum basiconicum smooth-surfaced sensillum basiconicum sensillum chaeticum corrugated surface cuticle dendrite pore of dermal gland dorsal surface dendritic protrusion dendritic sheath fusion of dendritic sheath with peg wall groove inner surface microvilli of trichogen cell outer surface pore canal pore plate pore tubule radial channel socket spatulatesensillumchaeticum sensillum fluid (Figs. 33, 34; 39); suspensory filaments (Fig. 28) dense strand (Figs. 40, 42); spatulate tip (Fig. 12) sensillum trichodeum tubular body trichogen cell trichogen cell vacuole tormogen cell ventral surface