Characterization of pericellular [ 125I]tyr0 dTrp8 somatostatin binding sites in the rat arcuate nucleus by a newly developed method: Quantitative high-resolution light microscopic radioautography

Abstract

In the present work we characterized the kinetic properties of [ 125I]somatostatin pericellular binding sites in the arcuate nucleus of the hypothalamus of the rat by quantitative high-resolution light microscopic radioautography. In order to determine whether these pericellular binding sites corresponded to functional receptors, their properties were compared with those of previously well-characterized [ 125I]somatostatin binding sites present on neuronal processes on the same sections in the stratum radiatum of the CA1 of the hippocampus. Radiolabelled sections were analysed by densitometry using a Biocom image analysis system coupled with a Leitz orthoplan microscope. The linear relationship between optical densities and radioactive standards allowed us to quantitate [ 125I]somatostatin-specific binding. Binding was time- and temperature-dependent, and saturable and specific in the arcuate nucleus as in the CA1 of the hippocampus. Saturation experiments indicated a single receptor population of binding sites with K D values of 0.2 ± 0.1nM in the arcuate nucleus and 0.6 ± 0.4nM in the CA1. In both structures, displacement curves obtained with somatostatin 14 and somatostatin 28 were monophasic, but shallow, while the somatostatin analogue SMS 201–995 induced a biphasic displacement, suggesting two populations of binding sites. In both regions binding was GTP-dependent. Desaturation procedures ( in vivo by cysteamine and in vitro by preincubating with GTP) resulted in an increase in the number of measurable binding sites. This increase was more marked in the arcuate nucleus than in the CA1, suggesting that the rate of occupancy of the binding sites by endogenous somatostatin is higher in the arcuate nucleus than in the CA1. This last observation is suggestive of a dynamic role of somatostatin: i.e. regulating a subpopulation of growth hormone-releasing factor-containing arcuate neurons.