In the research group we are interested in the development of the antennal lobes of the sphinx moth Manduca sexta.

The olfactory system of insects consists of three major components.

1. The Antenna:

Here the olfactory sensilla with the olfactory receptors are located.

2. The antennal lobes:

Spherical structures anteriorly located on both sides of the brain - they are also called deutocerebrum. In the antennal lobe - depending on the insect species - a variable number of small spherical structures (glomeruli) and neurons can be found. The axons of the antennal receptor cells and the the neurons of the antennal lobe both project in the glomeruli to build a dense meshwork of synapses. Thus the antennal lobe is the first integration center for odor information. Glomeruli can be found in all first odor integration centers in the animal kingdom, like for example in our olfactory bulb, the vertebrate analogue to the insects antennal lobe. Because of this morphological similarity it is assumed, that signal processing is in principal similarly working within different animal phyla. Recent functional studies using calcium imaging underline this idea. The experiments show the glomeruli in insects and in vertebrates as functional units, in which the various odor combinations are odotopical represented.

3. Higher integration centers:

From the antennal lobe neurons project to other parts of the central brain, like the mushroom bodies or the lateral protocerebrum. Here, the odor signal is further processed. The mushroom bodies for example are necessary for insects to obtain odor memories.

The antennal lobe of the sphinx moth Manduca sexta is a well acknowledged model system in terms of studying the development of the olfactory system. In holometabolous insects like M. sexta the antennal lobe is built during metamorphosis (=adult development or pupal development). The larva of M. sexta has only a few neurons in a so called larval antennal center to process the olfactory information from about 10 sensilla on the larval antenna. Adult M. sexta males have about 400.000 olfactory receptor cells on one antenna, which carry information about the odor world into the central brain. In order to process this enormous amount of information a structure like the antennal lobe is built during metamorphosis.

Ms_larva_pupa_adult_transparent

Since today we do not know how the neuronal network in the antennal lobe is acquired, how the different types of neurons find their partners, how the synaptic strength between the neuronal partners is determined.

Projects

In the group we are mainly interested in two aspects of antennal lobe development:

1. Regulation and role of NO (nitric oxide) and cGMP (cyclic guanylyl monophosphate)

We were able to demonstrate that (1) during metamorphosis about hundred local interneurons (about 10 % of the neuronal population of the lobe) use the nitric oxide NO/cGMP signaling pathway, (2) that the ability to use this pathway is developmentally regulated by the steroidhormone 20-hydroxyecdysone and (3) that the usage of the pathway depends on activity of the olfactory receptor cells from the antenna. (4) Furthermore, biogenic amines seem to be involved in cGMP regulation. The developmental window in which the neurons are sensitive to NO parallels the wave of synaptogenesis in the antennal lobe. This parallel together with results from work in vertebrates as well as in insects suggests a role of the NO/cGMP signaling pathway during synaptogenesis.

This project is since June 1999 supported by a grant from the DFG (Scha 678/3-1and Scha 678/3-3).

2. Regulation of peptide-identities of antennal lobe neurons and which role these peptides may play during antennal lobe development.

In adult M. sexta a large number of peptides is expressed in neurons of the antennal lobe. It is nothing known about which role these substances play during signal transmission. Also nothing is known about when during development these neurons acquire their peptide identities nor whether these peptides play an active role during development.

So far we examined the expression pattern of various peptides during antennal lobe development. We found that the developmental expression patterns are regulated via 20-hydroxyecdysone. For example increases the number of -RFamide positive neurons with increasing 20-hydroxyecdysone hemolymph levels, stops then at a certain hormone level to increase again with a falling hormone titer. In contrast the number of allatostatin positive neurons steadily increases with increasing hormone levels.

Methods

In our work we use various techniques like - pharmacological manipulation, immunocytochemistry on the light- and electronmicroscopic level, confocal laserscan microscopy, Westernblot technique, ELISA, electrophysiology, MALDI-TOF, Ca²⁺ - Imaging.

Selected Literature

Anton S, Homberg U (1999) Antennal lobe structure. In: Insect Olfaction. B. S. Hansson, Ed. Springer Verlag, Berlin, Heidelberg, New York, 97-124.
Ball EE , Truman JW (1998) Developing grasshopper neurons show variable levels of guanylyl cyclase activity on arrival at their targets. J Comp Neurol 394:1-13.
Bicker G (1998). NO news from insect brains. TINS 21:349-355.
Dubuque SH, Schachtner J, Nighorn AJ, Menon K, Zinn K, Tolbert LP (2001) Immunolocalization of synaptotagmin for the study of synapses in the developing antennal lobe of Manduca sexta. J Comp Neurol 441:277-287.
Gibbs SM , Truman JW (1998). Nitric oxide and cyclic GMP regulate retinal patterning in the optic lobe of Drosophila. Neuron 20:83-93.
Gibbs SM , Becker A, Hardy RW, Truman JW (2001) Soluble guanylate cyclase is required during development for visual system function in Drosophila. J Neurosci 21:7705-7714.
Gibson NJ, Rössler W, Nighorn AJ, Oland LA, Hildebrand JG, Tolbert LP (2001) Neuron-glia communication via nitric oxide is essential in establishing antennal-lobe structure in Manduca sexta. Dev Biol. 2001 Dec 15;240(2):326-39.
Higgins MR, Gibson NJ, Eckholdt PA, Nighorn A, Copenhaver PF, Nardi J, Tolbert LP (2002) Different isoforms of fasciclin II are expressed by a subset of developing olfactory receptor neurons and by olfactory-nerve glial cells during formation of glomeruli in the moth Manduca sexta. Dev Biol. 244:134-54.
Hildebrand JG, Rössler W, Tolbert LP (1997) Postembryonic development of the olfactory system in the moth Manduca sexta: primary-afferent control of glomerular development. Cell Dev Biol 8:163-170.
Hildebrand JG , Shepherd GM (1997) Mechanisms of olfactory discrimination: converging evidence for common principles across phyla. Annu Rev Neurosci 20:595-631.
Homberg U, Müller U (1999) Neuroactive substances in the antennal lobe. In: Insect Olfaction. B. S. Hansson, Ed. Springer Verlag, Berlin, Heidelberg, New York, 181-206.
Kaneko M, Nighorn A (2003) Interaxonal Eph-ephrin signaling may mediate sorting of olfactory sensory axons in Manduca sexta.
J Neurosci. 23:11523-38.
Morton DB, Nighorn A. (2003) MsGC-II, a receptor guanylyl cyclase isolated from the CNS of Manduca sexta that is inhibited by calcium.
J Neurochem. 84:363-72.
Müller U (1997) The nitric oxide system in insects. Progr Neurobiol 51:363-381.
Nighorn A , Gibson NJ, Rivers DM, Hildebrand JG, Morton DB (1998) The nitric oxide-cGMP pathway may mediate communication between sensory afferents and projection neurons in the antennal lobe of Manduca sexta. J Neurosci 18:7244-7255.
Nighorn A , Gibson NJ (2000) Expression of nitric oxide synthase and soluble guanylyl cyclase in the developing olfactory system of Manduca sexta. J Comp Neurol.422:191-205.
Oland LA , Tolbert LP (1996). Multiple factors shape development of olfactory glomeruli: insights from an insect model system. J Neurobiol 30:92-109.
Schachtner J , Klaassen L, Truman JW (1998) Metamorphic control of cyclic guanosine monophosphate expression in the nervous system of the tobacco hornworm, Manduca sexta. J Comp Neurol 396:238-252.
Schachtner J, Homberg U, Truman JW (1999) Regulation of cyclic GMP elevation in the developing antennal lobe of the sphinx moth, Manduca sexta. J. Neurobiol. 41:41:359-375.
Schachtner J, Huetteroth W, Nighorn A, Honegger HW (2004) Copper/Zinc superoxide dismutase-like immunoreactivity in the metamorphosing brain of the sphinx moth Manduca sexta. J Comp Neurol 469:141-152.
Truman JW , De Vente J, Ball EE (1996) Nitric oxide-sensitive guanylate cyclase activity is associated with the maturational phase of neuronal development in insects. Development 122:3949-3958.
Zayas RM , Qazi S, Morton DB, Trimmer BA (2000) Neurons involved in nitric oxide-mediated cGMP signaling in the tobacco hornworm, Manduca sexta. J Comp Neurol. 419:422-38.