Neuromuscular transmitter candidates of a centipede (Lithobius forficatus, Chilopoda).

BACKGROUND: The neuromuscular junction is the chemical synapse where motor neurons communicate with skeletal muscle fibers. Whereas vertebrates and many invertebrates use acetylcholine as transmitter at the neuromuscular junction, in those arthropods examined up to now, glutamate and GABA are used instead. With respect to taxon sampling in a phylogenetic context, there is, however, only a limited amount of data available, focusing mainly on crustaceans and hexapods, and neglecting other, arthropod groups. Here we investigate the neurotransmitter equipment of neuromuscular synapses of a myriapod, Lithobius forficatus, using immunofluorescence and histochemical staining methods. RESULTS: Glutamate and GABA could be found colocalised with synapsin in synaptic boutons of body wall and leg muscles of Lithobius forficatus. Acetylcholinesterase activity as a marker for cholinergic synapses was found abundantly in the central nervous system and also in some peripheral nerves, but not at neuromuscular junctions. Furthermore, a large number of leg sensory neurons displayed GABA-immunofluorescence and was also labeled with an antiserum against the GABA-synthesizing enzyme, glutamate decarboxylase. CONCLUSIONS: Our data indicate that glutamate and GABA are neurotransmitters at Lithobius forficatus neuromuscular junctions, whereas acetylcholine is very unlikely to play a role here. This is in line with the concept of glutamate as excitatory and GABA as the main inhibitory neuromuscular transmitters in euarthropods. Furthermore, we have, to our knowledge for the first time, localized GABA in euarthropod leg sensory neurons, indicating the possibility that neurotransmitter panel in arthropod sensory systems may be far more extensive than hitherto assumed.

Regeneration of synapses in the olfactory pathway of locusts after antennal deafferentation.

The olfactory pathway of the locust is capable of fast and precise regeneration on an anatomical level. Following deafferentation of the antenna either of young adult locusts, or of fifth instar nymphs, severed olfactory receptor neurons (ORNs) reinnervate the antennal lobe (AL) and arborize in AL microglomeruli. In the present study we tested whether these regenerated fibers establish functional synapses again. Intracellular recordings from AL projection neurons revealed that the first few odor stimulus evoked postsynaptic responses from regenerated ORNs from day 4-7 post crush on. On average, synaptic connections of regenerated afferents appeared faster in younger locusts operated as fifth instar nymphs than in adults. The proportions of response categories (excitatory vs. inhibitory) changed during regeneration, but were back to normal within 21 days. Odor-evoked oscillating extracellular local field potentials (LFP) were recorded in the mushroom body. These responses, absent after antennal nerve crush, reappeared, in a few animals as soon as 4 days post crush. Odor-induced oscillation patterns were restored within 7 days post crush. Both intra- and extracellular recordings indicate the capability of the locust olfactory system to re-establish synaptic contacts in the antennal lobe after antennal nerve lesion.

Regeneration of axotomized olfactory neurons in young and adult locusts quantified by fasciclin I immunofluorescence.

The olfactory pathway of the locust Locusta migratoria is characterized by a multiglomerular innervation of the antennal lobe (AL) by olfactory receptor neurons (ORNs). After crushing the antenna and thereby severing ORN axons, changes in the AL were monitored. First, volume changes were measured at different times post-crush with scanning laser optical tomography in 5th instar nymphs. AL volume decreased significantly to a minimum volume at 4 days post-crush, followed by an increase. Second, anterograde labeling was used to visualize details in the AL and antennal nerve (AN) during de- and regeneration. Within 24 h post-crush (hpc) the ORN fragments distal to the lesion degenerated. After 48 hpc, regenerating fibers grew through the crush site. In the AL, labeled ORN projections disappeared completely and reappeared after a few days. A weak topographic match between ORN origin on the antenna and the position of innervated glomeruli that was present in untreated controls did not reappear after regeneration. Third, the cell surface marker fasciclin I that is expressed in ORNs was used for quantifying purposes. Immunofluorescence was measured in the AL during de- and regeneration in adults and 5th instar nymphs: after a rapid but transient, decrease, it reappeared. Both processes happen faster in 5th instar nymphs than in adults.