Characterization of dopamine D2 receptors in the pituitary of the African catfish, Clarias gariepinus.

Dopamine receptors in the pituitary of the African catfish, Clarias gariepinus, were characterized using [3H]spiperone as radioligand. Specific binding of [3H]spiperone to pituitary membranes reached equilibrium within 60 min of incubation. The binding of the radioligand was tissue specific since the amount of binding was linear with pituitary membrane content in the incubations. In addition, pituitary membranes were observed to bind considerably more [3H]spiperone, compared to membrane preparation of various other tissues. Saturation experiments revealed the presence of a single class of high affinity/low capacity binding sites. The binding characteristics, estimated by Scatchard analysis, were: Kd = 3.2 +/- 0.5 x 10(-9) M and Bmax = 105 +/- 5 fmol/mg protein. Specific binding was displaceable with dopamine and with various specific D2 agonists and antagonists. The nature of displacement curves resembles those observed in studies on mammalian dopamine receptors. Binding experiments with cell fractions, obtained after centrifugation of dispersed pituitary cells over a Percoll density gradient, showed that most [3H]spiperone binding was obtained in an enriched gonadotropic cell fraction. This observation indicates that the receptor characteristics, estimated with the [3H]spiperone assay, are representative for dopamine receptors on the gonadotropic cells.

Role of calcium ions in action of gonadotropin-releasing hormone on gonadotropin secretion in the African catfish, Clarias gariepinus.

The aim of the present study was to establish the role of calcium ions in the mechanism of action of gonadotropin-releasing hormone (GnRH) in stimulating gonadotropin (GTH) release in the African catfish, Clarias gariepinus. For that purpose, GTH release from pituitary fragments was monitored in a perifusion system. GTH release, induced by the GnRH analog Buserelin, was strongly diminished in the absence of Ca2+, as well as in the presence of the Ca2+ channel antagonist nifedipine. In addition, the Ca2+ ionophore A23187 stimulated GTH secretion in the absence of GnRH. These results indicate that calcium ions play an intermediate role in the mechanism of action of GnRH in the African catfish.

Catecholestrogens inhibit dopamine methylation in the gonadotrops of the African catfish,Clarias gariepinus.

Isolated gonadotrops of the African catfish,Clarias gariepinus, were incubated with dopamine (DA) and/or catecholestrone and the activity of the enzyme catechol-O-methyltransferase (COMT) was determined by measuring the methylated products. From the apparent Km values for DA and catecholestrone of 0.4-1.3 µM and 17.9-25.2 µM respectively, it was concluded that catecholestrone is a better substrate for the enzyme COMT, compared to DA. Moreover, the methylation of DA is inhibited by comparatively low concentrations of catecholestrone.

Immunocytochemical evidence for peptidergic (GnRH) and dopaminergic innervation of the gonadotropic cells in the pituitary of the African catfish, Clarias gariepinus.

The proximal pars distalis (PPD) of the pituitary of the African catfish, Clarias gariepinus, was studied with immunocytochemical methods at the ultrastructural level. Anti-serum raised against synthetic mammalian luteinizing hormone-releasing hormone (LHRH) was applied on Lowicryl-embedded pituitaries and the antigenic sites were visualized with protein A-gold. In nerve fibers contacting the gonadotropic cells, granulated vesicles with a diameter of 90-120 nm were labeled after this procedure, whereas the glandular cells were not labeled. For the immunocytochemical demonstration of dopaminergic fibers, the preembedding method was performed on Vibratome sections, using highly specific antibodies against dopamine. Immunoreactivity was restricted to fibers containing granulated vesicles with a diameter of approximately 80 nm and terminating on gonadotropic cells. The present data support the results of earlier in vivo and in vitro studies on the catfish pituitary, indicating a dual neuroendocrine regulation of the gonadotropic cells.

Specificity and memory in increased defence reactions against bacteria in the pond snail Lymnaea stagnalis.

In Lymnaea stagnalis injections with dead Escherichia coli or Staphylococcus saprophyticus bacteria resulted in an enhanced clearance of both live S. saprophyticus and E. coli injected 4 days later. A non-specific activation of the internal defence system was concluded from these findings. The activation was dose-dependent: pre-injections with high doses resulted in a higher increase in the clearance capacity of the snails than pre-injections with low doses of bacteria. The state of increased activity of the defence system, induced with injection of dead E. coli, lasted at least 64 days. The heightened responses of the defence system were probably due to an activation of the blood cells (amoebocytes) since: 1) amoebocyte numbers increased faster in bacteria pretreated snails than in control animals; 2) ultrastructural observations revealed that the amoebocytes of bacteria pre-treated animals had a more ruffled outline than those of control snails; 3) amoebocytes from sensitized snails showed a higher phagocytic activity in vitro; 4) mitotic activity of amoebocytes increased after snails had been injected with bacteria.