Nitric oxide as a putative messenger molecule in the crayfish olfactory midbrain.

NADPH-d histochemistry was used to investigate presumptive nitric oxide synthase (NOS)-containing neurons in the crayfish olfactory midbrain. Three anatomically different types of local olfactory interneurons exhibiting NADPH-d activity were observed: two pairs of large interneurons as well as positively stained globuli cells. Branches derived from the large interneurons were confined to the ipsilateral olfactory lobe and accessory lobe, but only a few branches innervated the olfactory lobe glomeruli. Local field potential recordings on the olfactory lobe showed that administration of SNP or SIN-1 (10-4 M) into the brain had reversible inhibitory effects on electrically-evoked responses of unidentified neuronal cell populations.

NO-synthase: what can research on invertebrates add to what is already known?

The present study attempts to review presently known data regarding the distribution of nitric oxide (NO) synthase and the function of NO in invertebrate species. NO is synthesized from L-arginine by the enzyme NO-synthase, and activates guanylate cyclase which in turn leads to an increase in levels of cGMP in target cells. Major contributions to the knowledge of NO as a messenger molecule in invertebrates have been made by NADPH-diaphorase histochemistry and biochemical assays. These techniques suggest the presence of a L-arginine/NO pathway in a variety of tissues, thus implicating multiple roles for NO in invertebrates.

NADPH-diaphorase histochemistry and nitric oxide synthase activity in deutocerebrum of the crayfish, Pacifastacus leniusculus (Crustacea, Decapoda).

The activity of an nitric oxide synthase in the deutocerebrum of the crayfish Pacifastacus leniusculus was investigated with histochemical and biochemical methods. By using the NADPH-diaphorase histochemical reaction, known as a selective marker for NO synthase in mammals, it was possible to localize specific neuronal elements in the crayfish. Pronounced diaphorase-staining was observed in peripheral olfactory sensory cells and in the neuropil of the olfactory lobes. Less intense diaphorase-staining also occurred in other deutocerebral neuropils, such as the accessory lobes, the lateral antennular neuropil and in the deutocerebral commissure neuropil. The biochemical assay revealed a calcium/calmodulin-dependent formation of citrulline from L-arginine in brain homogenate. It was also possible to show that the selective NO synthase inhibitor L-NOARG decreased the formation of citrulline. These data indicate a role for NO as an intercellular messenger in the crayfish.

The aesthetasc concept: structural variations of putative olfactory receptor cell complexes in Crustacea.

The structure of the aesthetascs has been investigated in the prawn Macrobrachium rosenbergii (larvae and juveniles), the opossum shrimp Neomysis integer, the euphausid Meganyctiphanes, and in the water-fleas Daphnia magna and D. longispina. The aesthetascs, that are thought to represent olfactory receptors, exhibit a considerable structural variation, ranging from the well known aesthetascs of higher crustaceans (lobster, crab, crayfish) to the corresponding sensilla found in the water-fleas and the males of opossum shrimps. The two following morphological characteristics of the aesthetascs are thought to indicate an olfactory function: the shape of the cuticular hair that is long and essentially hose-shaped, and the thin, loosely arranged cuticle of at least the outer part of the cuticular hair. The presence of other structural elements such as sensory cells, cilia, and enveloping cells are vital for the olfactory function, but the development is variable, which makes their use in the morphological definition of aesthetascs problematic.

Dopamine-immunoreactive neurons in the blowfly visual system: light and electron microscopic immunocytochemistry.

Dopamine-immunoreactive (DA-IR) neurons were mapped in detail in the visual system of the blowfly, Calliphora erythrocephala. Three types of DA-IR neurons could be identified in the optic lobes. One type constitutes a population of several thousand columnar small field amacrine neurons in the second neuropil region, the medulla. The other two types are large field projection neurons innervating the next, more central, synaptic region comprising the lobula and the lobula plate, as well as centres of the midbrain. Their cell bodies are located latero-ventrally in the brain. No DA-IR neurons were seen in the most peripheral visual synaptic neuropil, the lamina. The two types of projection neurons form overlapping wide field arborizations in the lobula and lobula plate and cannot be distinguished from each other in this region. Their central connections are different, however. One type of projection neuron, BOD1, consists of two neurons that bilaterally connect the optic lobes and neuropil on each side of the oesophageal foramen in the posterior protocerebrum. The other type, BOD2, also consists of two bilateral neurons similar to BOD1, but with their central processes posteriorly in the lateral protocerebrum. The amacrine DA-IR neurons form lateral processes in three layers of the medulla synaptic neuropil. These neurons were also investigated by means of electron microscopical immunocytochemistry. They contain predominantly clear vesicles, but a few dense core vesicles could be resolved. The synaptic connections of the DA-IR amacrines suggest that they form centrifugal feedback circuits between the inner and the outer portion of the medulla. The present results indicate that dopamine may be a neurotransmitter in functionally different classes of neurons of the blowfly visual system: amacrines and projection neurons.

Light and electron microscopic immunocytochemistry of neurons in the blowfly optic lobe reacting with antisera to RFamide and FMRFamide.

Different antisera to the molluscan cardioexcitatory peptide FMRFamide, and its fragment, RFamide (Arg-Phe-NH2), label a distinct population of neurons in the optic lobe of the blowfly, Calliphora erythrocephala. Seven morphological types of RFamide/FMRFamide-like immunoreactive neurons could be distinguished in the optic lobes based on the locations of their cell bodies, their axonal projections and the distribution of their processes. Of these, two types could be resolved in their entire extent, the others were labeled only in their cell bodies and terminal processes or were partly obscured by other immunoreactive processes. The RF-like immunoreactive neurons in the optic lobes are of two main classes: (1) two types of large field projection neurons and (2) five types of local neurons. One type of projection neurons (five in each lobe) connects the entire projected retinal mosaic of the medulla and lobula in the optic lobe with protocerebral centres associated with the mushroom body calyx. The other type (2-3 invading each lobe) has cell bodies in the protocerebrum and contralateral processes invading optic lobes. Of the class of local neurons there are two amacrine RF-like immunoreactive neurons in each medulla. Each of these amacrines supplies the entire mosaic with fine processes. The remaining local RF-like immunoreactive neurons are present in relatively large numbers (one type in more than 2000 copies in each medulla) and-supply the medulla, lobula and lobula plate neuropils with fine varicose processes. In the medulla the RF-like immunoreactive processes are arranged in strict layers whereas in the lobula complex the distribution is diffuse. Electron microscopic immunocytochemistry, using both pre-embedding immuno peroxidase-antiperoxidase and post-embedding protein A-gold labeling, was employed for analysis of cytology and synaptic connections of RF-like immunoreactive neurons in the medulla. The varicosities of the processes of the large field projection neurons were not found to make chemical synapses with other neurons in the medulla. The spines of the RF-like immunoreactive processes of the large medulla amacrines, however, make pre- and postsynaptic contacts with other neural elements. Our findings indicate that an RFamide/FMRFamide-like substance may be used as a neurotransmitter or neuromodulator by optic lobe neurons of different types. The local and projection RF-like immunoreactive pathways probably play different roles in visual processing.