PHARMACOLOGICAL CHARACTERIZATION OF SOMATOSTATIN RECEPTORS IN THE RAT CEREBELLUM DURING DEVELOPMENT

Abstract: Somatostatin (SRIF) receptors (SRIF‐Rs) are transiently expressed in a germinative lamina of the rat cerebellum, the external granule cell layer. The appearance of SRIF‐Rs coincides with the expression of SRIF‐like immunoreactivity in the cerebellum. However, the cellular location of SRIF‐Rs does not overlap with the distribution of SRIF‐like immunoreactivity, with the latter being restricted to ascending fibers arising from the brainstem, to perikarya within the white matter, and to some Purkinje cells. The characterization of SRIF‐Rs in the immature (13–day‐old) rat cerebellum was conducted by means of binding experiments in membraneenriched preparations and autoradiography, using two radioligands, [125I‐Tyr0,D‐Trp8]SRIF‐14 ([125I‐Tyr0,d‐Trp8]S14) and I25I‐SMS 204–090. The pharmacological profile of cerebellar SRIF‐Rs was compared with that of adult cortical SRIF‐Rs. Saturation studies performed in 13–day‐old rat cerebellum showed that the A'D values for [125I‐Tyr0,D‐Trp8]S14 and 125I‐SMS 204–090 binding were 0.35 ± 0.04 and 0.39 ± 0.01 nM, respectively. The corresponding Bmax values were 52.7 ± 4.8 and 49.9 ± 5.3 fmol/mg of protein, a result indicating that radioligands with high specific radioactivity (2,000 Ci/mmol) bind to a single class of high‐affinity sites (SSI). Competition studies showed that different D‐Trp‐sub‐stituted analogs displaced [125I‐Tyr0,d‐Trp8]S14 binding with Hill coefficients >1, a finding indicating the existence of different subtypes of binding sites. When [Tyr0,d‐Trp8]S14 was used as a competitor, two sites were resolved by Scatchard analysis in both 13–day‐old cerebellum and adult cerebral cortex. The higher‐affinity sites correspond to the SSI subtype identified in saturation experiments, whereas the lower‐affinity sites most likely correspond to the SS2 subtype. Ionic supplementation studies showed that divalent cations were required to obtain maximal specific binding on the SSI sites. In particular, Mn2+ was the most efficient cation for promoting binding of [125I‐Tyr0,d‐Trp8]S14. Addition of GTP to the incubation buffer induced a marked reduction of specific binding. The results obtained by membrane binding assays were similar to those obtained by quantitative autoradiography, a result indicating that the microenvironment of SRIF‐Rs was preserved in both types of tissue preparations. Receptors expressed in the developing rat cerebellum exhibited the same KD and similar pharmacological profile as those observed in the adult rat cortex. These results show that SRIF‐binding sites transiently expressed in the external granule cell layer of the cerebellum of young rats are indistinguishable from adult rat brain SRIF‐Rs. The extremely high density of SRIF‐Rs found in the external granule cell layer in 13–day‐old rats suggests that SRIF may play a pivotal role in the proliferation and/or differentiation of these germinative cells. Copyright © 1990, Wiley Blackwell. All rights reserved