Ground plan of the insect mushroom body: functional and evolutionary implications

Abstract

In most insects with olfactory glomeruli, each side of the brain possesses a mushroom body equipped with calyces supplied by olfactory projection neurons. Kenyon cells providing dendrites to the calyces supply a pedunculus and lobes divided into subdivisions supplying outputs to other brain areas. It is with reference to these components that most functional studies are interpreted. However, mushroom body structures are diverse, adapted to different ecologies, and likely to serve various functions. In insects whose derived life styles preclude the detection of airborne odorants, there is a loss of the antennal lobes and attenuation or loss of the calyces. Such taxa retain mushroom body lobes that are as elaborate as those of mushroom bodies equipped with calyces. Antennal lobe loss and calycal regression also typify taxa with short nonfeeding adults, in which olfaction is redundant. Examples are cicadas and mayflies, the latter representing the most basal lineage of winged insects. Mushroom bodies of another basal taxon, the Odonata, possess a remnant calyx that may reflect the visual ecology of this group. That mushroom bodies persist in brains of secondarily anosmic insects suggests that they play roles in higher functions other than olfaction. Mushroom bodies are not ubiquitous: the most basal living insects, the wingless Archaeognatha, possess glomerular antennal lobes but lack mushroom bodies, suggesting that the ability to process airborne odorants preceded the acquisition of mushroom bodies. Archaeognathan brains are like those of higher malacostracans, which lack mushroom bodies but have elaborate olfactory centers laterally in the brain.

Figure 1

Summary diagram of the principal elements of mushroom bodies in dicondylic insects. Top: Mushroom bodies with paired calyces (Ca) equipped with Kenyon cells from cell bodies known as globuli cells (gl). Kenyon cells (magenta) have dendritic branches in the calyces, which also receive inputs from sensory neuropils (green): diagrammed here, afferents from the antennal lobes that terminate laterally outside the calyx in the lateral horn neuropil of the protocerebrum (L ho). Afferents and efferents (aff, eff; brown) also supply and extend from various locations in the lobes (see Strausfeld, 2002). Lower: Calyxless mushroom body with globuli cells (gl) providing Kenyon cells (Kc) with axon-like branches to the vertical and median lobes (V, M). Inputs (afferent; aff) and outputs (efferents: eff) extend to and from various locations along the lobes.

Figure 2

Mushroom bodies of terrestrial Coleoptera. A-H. Reconstruction (A) from successive reduced-silver-stained sections (B-D, F-H) of the mushroom body of the soldier beetle Chauliognathus lecontei (E). I. Mushroom body of the fig beetle Cotinus mutabilis (J) labeled with anti-DCO to resolve Kenyon cells (magenta) and phalloidin (green), which reveals microglomeruli of the calyces (Ca). In beetles possessing antennal lobes (Ant Lo), projection neurons supply ascending tracts to the calyces (Ca) through the antenno-cerebral tract (ACT, green in panels A-D, F-H). In Coleoptera, each calyx is supplied by two identical populations of globuli cells (gl). The two calyces supply axons to the pedunculus (Ped) and the medial (M) and vertical (V) lobes. In some species, such as Chauliognathus lecontei, the concentric organization of each calyx is represented through the length of the pedunculus and its lobes. K. One of the paired glomerular antennal lobes (Ant Lo) of Cotinis mutabilis with the striate mechanosensory neuropil dorsocaudal to it (Mech S). Other abbreviations: FB, fan-shaped body of the central complex. Scale bar B (applies also to C, D, F-H) = 50 μm, Scale bars I, K = 50 μm.

Figure 3

Vertical and medial lobes of comparable size in a terrestrial tiger beetle (A Cicindela ocellata) and aquatic sunburst and whirligig beetles (D Thermonectus marmoratus; F Dineutus sublineatus) beetles show no clear differences in terms of size relative to other brain areas. Large argyrophilic processes indicate “extrinsic” (afferent and efferent) neurons to and from the lobes. B,C and F,G. Golgi impregnations of afferent supply to the lobes of T. marmoratus and D. sublineatus. B. In T. marmoratus both the vertical lobe (cross-section in squared bracket) and medial lobe (C, bracketed) receive terminals (highlighted black arrows) from the protocerebrum. F,G. In D. sublineatus the vertical lobe (F) and medial lobe (G) receive inputs (highlighted black arrow) and provide outputs (open arrow). Arrowheads in F indicate Kenyon cell processes. Scale bars = 50 μm.

Figure 4

Mushroom bodies of insects with secondary anosmia or reduced olfaction. A. Golgi impregnation showing Kenyon cells of the diving beetle Thermonectus marmoratus (C) provide a loose arrangement of dendrites (arrowed) that extend into the surrounding protocerebral neuropil. DCO labeling (magenta, inset a1) shows two bundles of Kenyon cell neurites (arrowheads) from globuli cells projecting into the protocerebrum without providing a definable calyx or even a remnant that, as in Odonata, would be represented by microglomeruli (green, inset b1). B. Kenyon cells of the mushroom body of the banded demoiselle damselfly Calopteryx splendens (D) provide long (arrowed) and short (s, open arrows) dendrites; the long ones extend into the surrounding protocerebrum, whereas the short dendrites provide their claw-like specializations at the pedunculus. In the dragonfly Perithemis tenera four bundles of neurites (arrowheads in inset b1) originate from four sets of globuli cells (labeled pale magenta with DCO in b1). These provide microglomeruli (labeled green with phalloidin, inset b2) within and surrounding the head of the pedunculus, shown in cross section. Scale bars = 50 μm, except inset b2, where the scale bar = 20 μm.

Figure 5

Mushroom bodies of the milkweed bug Oncopeltus fasciatus (panel A) compared with aquatic and water-adapted hemipterans (panels B-H)lacking antennal lobes and calyces. A-C, F, G show DCO/phalloidin labeling, D, E, H are Bodian-stained. A. O. fasciatus mushroom body showing globuli cells (gl), calyx (Ca) and pedunculus (Ped) extending through the lateral protocerebrum (L Pr) and giving rise to a medial lobe (M) equipped with numerous ovoid swellings. B. Notonecta undulata. Distal part of the pedunculus of this backswimmer supplied by globuli cells. The absence of microglomeruli suggests complete loss of calyx. The diffuse fascicle of axons (bracketed) comprises the vestigial antennocerebral tract. C. N. undulata. Medial lobe of mushroom body showing many swollen outgrowths. This contrasts with the medial lobe of the water strider Aquarius remigis, shown in H, which terminates as a single bulb. D. Aquarius remigis. The vertical lobes of the wider strider have two divisions (1,2) flanked by a smaller lobe (asterisk). E. N. glauca. The vestigial antennocerebral tract (bracketed) stained by reduced silver. In this species the mushroom body is simple, arising from one population of globuli cells with a pedunculus ending in a system of ovoid swellings, as shown in N. undulata in panel C. The mushroom body of N. glauca is reminiscent of the ephemeropteran mushroom body shown in Fig. 5A. F. Diceroprocta semicincta. The pronounced vertical (V) and medial (M) lobe of the cicada mushroom body that lacks a calyx. G. Diceroprocta semicincta. Mushroom body globuli cells (gl), pedunculus (ped) and residual antennocerebral tract (bracketed). H. A. remigis. The large bulbous medial lobe of the water strider. Scale bars = 50 μm.

Figure 6

Comparisons of ephemeropteran (panels A, a1-a3) and zygentoman (panels B-D) mushroom bodies. A. In the mayfly Potamanthus luteus mushroom bodies are simple A single cluster of globuli cells (gl) provides a pedunculus (Ped) the head of which contains an assembly of microglomeruli (green in inset a2, which is a DCO/phalloidin preparation corresponding to the white-outlined box in A), indicative of an attenuated calyx. The pedunculus extends medially to terminate as a few discrete swellings (arrows, also inset a3, a DCO/phalloidin preparation that corresponds to the black-outlined box in A). There is no evidence of a discrete vertical lobe in this taxon. B. Thermobia domestica. Mushroom bodies of the Zygentoma (inset b1 Lepisma saccharina) have a small calyx (Ca) composed of two components: a dense central aggregate of microglomeruli (here revealed by DCO) flanked by larger ones (curved brackets). These are supplied by two streams of axons from the antennocerebral tract. The smaller of the two (brackets, left) carries axons from the lobus glomerulatus. The larger tract carries axons of antennal lobe projection neurons. Globuli cells (gl) are organized in two clusters. C, D. Ctenolepisma longicaudata (silverfish) possesses mushroom bodies equipped with a medial lobe from which protrude grape-like swellings called Trauben (T) and a prominent vertical lobe (V). Bodian stains also show the calyx (Ca) supplied by a substantial antennocerebral tract (bracket in D). EB, ellipsoid body. Scale bars: A, B, C, D = 50 μm; inset a2 = 25 μm.

Figure 7

Mushroom body of the dragonfly Libellula depressa. A. Reduced-silver section showing two of the four bundles of neurites (arrowed) arising from several thousand globuli cells and supplying the two necks (ne) of the pedunculus (ped). B. Top-down view showing the two populations of globuli cells, each supplying one half of the mushroom body. The two necks of the pedunculus (ne) are surrounded by a swathe of protocerebral neuropil (Pro). There is no evidence in silver stains of a discrete calyx, but antisera against phalloidin reveal microglomeruli (Fig. 3B, inset b2). C. Golgi impregnation of a few dozen Kenyon cells supplying one pedunculus neck and then extending through the pedunculus and medial lobe. D-F. Enlargements of boxed areas in C show details of Kenyon cell dendrites that extend into the protocerebrum from the distal pedunculus (D) or that arise at several levels along the length of the pedunculus and medial lobe (E, F). Scale bars: A-C = 50 μm.

Figure 8

Parallel divisions of the mushroom body lobes. In neopteran insects, here exemplified by Phaenicia serricata (panels B, C, E) mushroom bodies consist of several discrete parallel divisions. Antisera against aspartate, taurine, and DCO distinguish divisions of the vertical and medial lobes (B, C, E). In odonate insects, here exemplified by the dragonfly Aeschna, the same terms (α, α′, β, β′, γ) can be used to indicate comparable subdivisions in the lobes (A, D, F inset, H). In the fly, intense DCO immunoreactivity identifies the lobelet (E), whereas in Aeschna an intense DCO-positive structure at the bulb of the vertical lobe (D) indicates a vertical branch of the γ-lobe. As in the fly (B), aspartate reveals two parallel divisions in the odonate mushroom body, best seen in cross sections of the pedunculus (inset to F). Aspartate labels the head of the odonate pedunculus (bracketed F) and the fly calyx (G). Taurine identifies the α′ division in the fly (C) and a comparable swelling at the tip of the odonate vertical lobe (asterisk in H). Scale bars = 50 μm.

Figure 9

Parallel divisions of the mushroom body of the odonate Libellula depressa. A. Cross sections of the pedunculus distinguish three parallel divisions, each of which will provide the corresponding components of the vertical (α, α′) and medial (β, β′) lobes with the γ division contributing to both. Note that each division has its own characteristic fibroarchitecture and extrinsic (input and output) elements. B. Afferent neurons invading the base of the pedunculus. C. The junction of the vertical and medial lobes. Afferent neurons extending to the α′/β′ (double arrows) shown with the initial branches of two large efferent neurons arising from the α/β and γ divisions (arrows). Scale bars = 50 μm.

Figure 10

Parallel and local organization of the lobes of Libellula depressa. A. Golgi impregnations demonstrate that at specific locations along the lobes Kenyon cells provide collaterals (bracketed) indicative of regions in which they participate in local circuits. B, C. Reduced silver impregnations reveal the participation of efferent (single black arrow in B, white arrows in C) and afferent (double (white and black arrows in B) neurons at the bulbous ending of the vertical (α) lobe. The spur (sp) indicates the peduncular divide at which Kenyon cells divide to send collaterals into the medial and vertical lobes. Scale bars = 50 μm.

Figure 11

An insect without a mushroom body: the archaeognathan Machilis germanicus (A). B. DCO/phalloidin shows no evidence of a mushroom body in the archaeognathan brain. However, there is a clearly delineated superior lateral protocerebrum (S L Pr), the organization of which is reminiscent of the medulla terminalis of a malacostracan crustacean, or the lateral horn olfactory neuropil of other insects. C. The machilid antenna provides discrete olfactory glomeruli (olf glom) and a more caudal antennal mechanosensory neuropil (ant mech). Gustatory neuropil (gust) is also glomerular and is situated in the tritocerebrum. Note the simple layered arrangement of the central complex (CC), again reminiscent of that of malacostracan crustaceans. Other abbreviations: Me, medulla; Lo, lobula; S M Pr, superior medial protocerebrum. Scale bars = 50 μm.

Figure 12

Comparisons of mushroom bodies in insects lacking a calyx and antennal lobes with an insect (Cicindela ocellata) equipped with both. Mushroom body neuropils are shown green, and globuli cell clusters supplying them are magenta. Of the six midbrains shown here, only that of the tiger beetle Cicindela ocellata possesses antennal lobes (orange) and an ascending tract supplying calyces. The mushroom body of basal palaeopterans (exemplified by the damsel fly Agrion virgo, and mayfly Potamanthus luteus) possesses lobes, those of the odonate are substantial. Secondarily aquatic neopterans, here the sunburst beetle Thermonectus marmoratus and the hemipteran Notonecta undulata (backswimmer) have calyxless mushroom bodies, the latter showing tuberous outgrowths from the base of the pedunculus that are reminiscent of those in Potamanthus (see Fig. 6A). The terrestrial homopteran, the cicada Diceroprocta semicincta, lacks antennal lobes but possesses calyxless mushroom bodies.

Figure 13

Similarities between flightless and winged dicondylian insects, here the zygentoman apterygote Lepisma saccharina (“silverfish”, left) compared with the orthopteran Schistocerca gregaria (“Rocky Mountain locust”, right). In both taxa mechanosensory terminals from the antennae (AN) supply the tritocerebrum, and olfactory receptors from the antennae supply axons to the antennal lobe in the deutocerebrum. In both, the lobus glomerulatus receives gustatory inputs and, like the antennal lobe, provides a separate stream of projection neuron axons to a discrete region of the calyx (Ca I, accessory calyx). Gustatory and olfactory projection neuron axons appear to extend beyond their specific calyx regions (Ca I, Ca II) into the lateral horn (LH) of the superior lateral protocerebrum. gc globuli cells; M medial lobe; V vertical lobe

Figure 14

Elaboration of the mushroom body ground plan as evidenced by the apterygote Zygentoma (upper left panel: calyces receive olfactory inputs (red)). In all mushroom bodies, with or without a calyx [the latter as in the Ephemeroptera (upper middle) and certain Hemiptera], parallel fibers contribute to local networks that receive inputs (purple) and provide outputs (black). In odonates (upper right), the distal part of the pedunculus reveals a remnant calyx equipped with microglomeruli receiving inputs (blue) from visual interneurons. Calyces are most elaborated when they serve several modalities, as in many species of Hymenoptera (olfactory input red, visual input blue). Modality-specific zones of the calyces supply longitudinally partitioned lobes (lower left). In some Neoptera, the entire calyx is dominated by a modality other than olfaction, as in the “visual” calyces of the Gyrinidae (lower middle). Irrespective of how many modalities supply the calyces, the lobes of most mushroom bodies are divided into parallel or concentric divisions (lower right). These are usually represented by a concentric organization of Kenyon cell populations in the calyces.

Figure 15

Evolution of mushroom bodies. Glomerular olfactory lobes supplied by the first antennae (character 1, see Note on Terminology) are resolved in malacostracan Crustacea and in the Archaeognatha. Projection neurons from the glomerular lobes extend to the lateral protocerebrum of both malacostracan crustaceans and archaeognathans (character 2). Mushroom bodies with calyces and lobes (3), supplied by olfactory glomeruli and gustatory inputs (4) are resolved in the Zygentoma and are thus basal to Palaeoptera and Neoptera. Double calyces (5) and parallel subdivisions of the lobes (6) are resolved in pterygote insects. The representation of modalities in addition to olfaction and taste (visual afferents (7) are resolved in the Odonata and higher Neoptera. Modality substitution (8; vision substituting olfaction) in the calyces or their rudiments has occurred independently in some Coleoptera and Odonata. Secondary loss of antennal lobe glomeruli and the reduction or complete loss of calyces (9, 10) has occurred several times independently within the Palaeoptera and Neoptera. Aquatic species are shown in bold type. The phylogeny on which characters 1-10 are mapped is based on Wheeler et al., 2001.