Evidence for the cholinergic markers ChAT and vAChT in sensory cells of the developing antennal nervous system of the desert locust Schistocerca gregaria

Sensory and motor systems in insects with hemimetabolous development must be ready to mediate adaptive behavior directly on hatching from the egg. For the desert locust S. gregaria, cholinergic transmission from antennal sensillae to olfactory or mechanosensory centers in the brain requires that choline acetyltransferase (ChAT) and the vesicular acetylcholine transporter (vAChT) already be present in sensory cells in the first instar. In this study, we used immunolabeling to demonstrate that ChAT and vAChT are both expressed in sensory cells from identifiable sensilla types in the immature antennal nervous system. We observed ChAT expression in dendrites, neurites and somata of putative basiconic-type sensillae at the first instar stage. We also detected vAChT in the sensory axons of these sensillae in a major antennal nerve tract. We then examined whether evidence for cholinergic transmission is present during embryogenesis. Immunolabeling confirms that vAChT is expressed in somata typical of campaniform sensillae, as well as in small sensory cell clusters typically associated with either a large basiconic or coeloconic sensilla, at 99% of embryogenesis. The vAChT is also expressed in the somata of these sensilla types in multiple antennal regions at 90% of embryogenesis, but not at earlier (70%) embryonic stages. Neuromodulators are known to appear late in embryogenesis in neurons of the locust central complex, and the cholinergic system of the antenna may also only reach maturity shortly before hatching.

Neuromodulation by 5-hydroxytryptamine in the antennal lobe of the sphinx moth Manduca sexta.

Using intracellular recording techniques, we have begun to examine the effects of 5-hydroxytryptamine (5-HT) on antennal-lobe (AL) neurones in the brain of the adult moth Manduca sexta. 5-HT modulated the responses of local interneurones and projection neurones, which were recognized on the basis of well-established electrophysiological criteria, to primary synaptic input elicited by electrical stimulation of the ipsilateral antennal nerve. 5-HT applied at low concentration (10(-8) mol l-1) reduced the excitatory responses evoked by electrical stimulation of the antennal nerve, whereas at high concentration (10(-4) mol l-1), 5-HT enhanced the responses. At 10(-4) mol l1, 5-HT increased cell input resistance, led to broadening of action potentials and caused increased cell excitability in many AL neurones.

The development of the sensory neuron pattern in the antennal disc of wild-type and mutant (lz3, ssa) Drosophila melanogaster

The development of the sensory neuron pattern in the antennal disc of Drosophila melanogaster was studied with a neuron-specific monoclonal antibody (22C10). In the wild type, the earliest neurons become visible 3 h after pupariation, much later than in other imaginal discs. They lie in the center of the disc and correspond to the neurons of the adult aristal sensillum. Their axons join the larval antennal nerve and seem to establish the first connection towards the brain. Later on, three clusters of neurons appear in the periphery of the disc. Two of them most likely give rise to the Johnston's organ in the second antennal segment. Neurons of the olfactory third antennal segment are formed only after eversion of the antennal disc (clusters t1-t3). The adult pattern of antennal neurons is established at about 27% of metamorphosis. In the mutant lozenge3 (lz3), which lacks basiconic antennal sensilla, cluster t3 fails to develop. This indicates that, in the wild type, a homogeneous group of basiconic sensilla is formed by cluster t3. The possible role of the lozenge gene in sensillar determination is discussed. The homeotic mutant spineless-aristapedia (ssa) transforms the arista into a leg-like tarsus. Unlike leg discs, neurons are missing in the larval antennal disc of ssa. However, the first neurons differentiate earlier than in normal antennal discs. Despite these changes, the pattern of afferents in the ectopic tarsus appears leg specific, whereas in the non-transformed antennal segments a normal antennal pattern is formed. This suggests that neither larval leg neurons nor early aristal neurons are essential for the outgrowth of subsequent afferents.

Particle arrays on insect nerve membranes

Freeze-fracture studies of Lepidopteran antennae have revealed the presence of orthogonal arrays of particles on antennal nerve membranes. Initial impressions were that several different arrays were present. Optical diffraction was used to examine the arrays on the electron micrograph negatives. This technique showed that all the arrays were derived from the same basic structure, suggesting that the superficial differences in appearance were due to shadowing effects. The particles are arranged in a lattice with spacings of 10.7 nm X 9.1 nm. The arrays are not clear-cut but tend to break up, producing a disorganized region around their edges. The particles are shown to have depressions in them. However, the evidence available suggests that the arrays do not have the other characteristics of gap junctions. The arrays appear not to be present on most of the nerve membrane faces, occurring only in localized regions of the nerve membranes where they are present in large numbers. This suggests that the arrays may have a specialized local function.

Organization and synaptic ultrastructure of glomeruli in the antennal lobes of the moth Manduca sexta : a study using thin sections and freeze-fracture

The antennal lobe of the brain of Manduca sexta comprises a central area of coarse neuropil surrounded by dense, spheroidal glomeruli, where all synaptic interactions between antennal-nerve axons and the second-order neurons of the lobe occur. Neuronal interactions in the glomeruli are complex, involving several types of neuritic profiles and mediated by synapses with a one-to-many ratio of pre- to postsynaptic elements. Presynaptic profiles in the glomeruli have been categorized into three types, containing round clear vesicles, large numbers of large dense-cored vesicles, and pleiomorphic clear vesicles, respectively. Preliminary studies of horseradish peroxidase-filled axons and neurons indicate that antennal-nerve axons form synapses without large numbers of dense-cored vesicles and that antennal-lobe neurons not only receive synapses but also may synapse onto other elements in the antennal lobe. A typical synaptic contact involves multiple postsynaptic elements apposed in pairs to an individual presynaptic element. The presynaptic element contains a bar-shaped membrane-associated density, which follows a shallow groove in the membrane and is flanked by synaptic vesicles. Postsynaptic elements are lined by membrane-associated densities in the region opposite to the synaptic bar, and may be observed to participate in serial synapses. Freeze-fracture replicas of the glomerular neuropil contain many membrane specializations that are thought to be presynaptic, some of which resemble those of vertebrate excitatory synapses. At these apparently presynaptic regions, large particles cluster in the P face of the membrane and are often surrounded by plasmalemmal deformations presumably representing sites of exo- or endocytosis. The shape of the predominant type of presynaptic membrane specialization (a plaque) does not match the shape of the presynaptic membrane-associated density (a bar); this raises the possibility that vesicle release occurs at isolated ‘active zones’ along the presynaptic bar. Postsynaptic sites are represented by clusters of large particles in the E face of the postsynaptic membrane.