The pharmacological profile and localization of somatostatin (SRIF) receptors were determined in rat, monkey and human cerebellum. In rat cerebellar cortex, low sst(1)/sst(4), intermediate sst(2) and very high sst(3) receptor mRNA levels were found. sst(1) mRNA was also expressed in the deep cerebellar nuclei [I-125]Tyr(3)-octreotide binding sites in cerebellar membranes correlated with recombinant sst(2), but not with sst(5) or sst(3) receptors and were found in the molecular layer of the cerebellum. [I-125]CGP 23996 (in Na+-buffer) binding in rat cerebellum correlated with sst(1) or sst(4), but not with sst(2), sst(3) or sst(5) receptor binding. Similar data were obtained in rhesus monkey cerebellum. mRNAs for all five receptors were found in the granule cell layer of the human cerebellum and/or in the dentate nucleus. [I-125]Tyr(3)-octreotide binding was strong in the molecular layer and correlated with that of recombinant sst(2) receptors, but not with sst(3) or sst(5) receptors. [I-125]CGP 23996 (in Mg++-buffer) binding was heterogeneous (about 75%. to sst(2) and 25% to sst(1) and/or sst(4) receptors). The molecular and granular layers were equally and the dentate nucleus strongly labeled. Thus, SRIF receptors of the sst(2), sst(1) and/or sst(4) subtype are present in the rat, monkey and human cerebellum. In the latter two species, the sst(2) type appears to be predominant. Surprisingly, the high expression of sst(3) receptor mRNA is not supported by radioligand binding data in any of the species studied. The reason for this discrepancy remains to be elucidated.

In situ hybridization histochemistry with somatostatin sst(1)-sst(5) receptor messenger RNA-selective oligoprobes and quantitative receptor autoradiographic binding studies using [I-125]Tyr(3)-octreotide, [Leu(8), D-Trp(22),I-125-Tyr(25)]somatostatin-28 and [I-125]CGP 23996 ([I-125]c[Asn-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Tyr-Thr-Ser]) were performed to determine the level of expression of somatostatin receptor messenger RNA and receptor binding sites in the hippocampal formation, limbic system and cerebral cortex of adult rats electrically kindled in the dorsal hippocampus.In control rats (implanted with electrodes but not electrically stimulated), the somatostatin-1 receptor-selective [I-125]Tyr(3)-octreotide and the non-subtype-selective [LeU(8),D-Trp(22),I-125-Tyr(25)]somatostatin-28 preferentially labelled the strata oriens and radiatum of the CA1 subfield of the hippocampus, the molecular layer of the dentate gyrus, the subiculum and presubiculum of the hippocampal formation, the inner layer of the frontal cortex, and the lateral and basolateral nuclei of the amygdala. The non-subtype-selective radioligand [I-125]CGP 23996 (in 5 mM Mg2+ buffer) preferentially labelled the strata oriens and radiatum of the CA1 subfield of the hippocampus, the subiculum and the basolateral nucleus of the amygdala. Under conditions where primarily somatostatin-2 receptors were labelled, [I-125]CGP 23996 (in 120 mM Na+ buffer) showed strong binding in the strata oriens and radiatum of the CA1 subfield of the hippocampus and the frontal cortex, whereas the dentate gyrus, subiculum and amygdala showed only weak signals. During and after kindling, no significant differences were observed between the ipsi- and contralateral sides of the hippocampus. A significant decrease (about 40%) of somatostatin receptor binding sites was observed in the molecular layer of the dentate gyrus with all radioligands (except [I-125]CGP 23996 in Na+ buffer, which did not label this area) at stage 2 (pre-convulsive stage) and one week, but not one month, after stage 5 (generalized motor seizures). In contrast to somatostatin receptor binding, no alterations of the messenger RNA levels for sst(1)-sst(5) receptors were found either at stage 2 or at stage 5. Similarly, no changes in receptor binding or messenger RNA levels were observed in the brain of rats which experienced a single afterdischarge.The present study shows a significant and selective decrease of somatostatin-1 receptor binding sites in the dentate gyrus of kindled rats. This is part of the plastic changes induced by kindling and may contribute to the increased sensitivity for the induction of generalized seizures during kindling. Copyright (C) 1996 IBRO.

Radioligand binding studies were performed in membranes of human cerebellum using [125I][Tyr3]octreotide also known as [125I]204-090, [125I]LTT-SRIF-28 ([Leu8, d-Trp22, 125I-Tyr25]SRIF-28) and [125I]CGP 23996 ([125I]c[Asu-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Tyr-Thr-Ser]) to characterize the nature of cerebellar somatostatin receptors. Saturation experiments performed with [125I]204-090 suggest the presence of a single class of binding sites with high affinity: Bmax = 55.7 ± 9.7 fmol/mg protein, pKd = 9.57 ± 0.04. The pharmacological profile of [125I]204-090 and [125I]LTT-SRIF-28 labelled sites in human cerebellar membranes was overlapping (correlation coefficient r = 0.998) and correlated very significantly with that of recombinant human sst2 receptors (r = 0.987). By contrast, there was very little correlation with those of recombinant human sst3 (r = 0.208) or human sst5 receptors (r = 0.547). In contrast to [125I]204-090 or [125I]LTT-SRIF-28 binding, [125I]CGP 23996 binding (in 5 mM MgCl2 buffer) in cerebellar membranes was heterogeneous as indicated by biphasic competition curves produced by sst2 receptor selective ligands such as seglitide or octreotide. The pharmacological profile of the major component was closely correlated with that of human sst2 receptors (r = 0.989), whereas the minor component correlated equally well with human sst1 or sst4 receptors (r = 0.902 and 0.941, respectively). In vitro autoradiographic studies performed in cerebellar slices using [125I]204-090 and [125I]LTT-SRIF-28 demonstrated the presence of binding sites predominantly in the molecular layer, whereas weaker labelling was detected in the granular layer. The distribution of sites labelled by both radioligands was very similar. Using [125I]CGP 23996 (in 120 mM NaCl buffer), no clear difference between labeling of the molecular and granular layers was detectable; the dentate nucleus demonstrated binding sites for [125I]CGP 23996, in contrast to the very low level of binding observed with both, [125I]204-090 and [125I]LTT-SRIF-28. Together, the present data demonstrate the presence of SRIF receptors in the adult human cerebellar cortex which are, for the major population, best characterized as sst2. The SRIF receptors in the minor population of the cerebellar cortex and the dentate nucleus most probably represent sst1 and/or sst4 sites.

A transient expression of somatostatin mRNA as well as of the peptide itself has been described in the developing mammalian auditory brainstem. However, little is known about the presence, and the spatial and temporal pattern of somatostatin (SRIF) receptor subtypes in this system. Therefore, we investigated the distribution of SRIF receptor binding sites labeled with the radioligands [125I]LTT-SRIF-28, [125I]Tyr3-octreotide, and [125I]CGP 23996 (in buffers containing either Mg2+ or Na+ ions) within the developing auditory brainstem of the rat. In addition, we performed in situ hybridization with a 33P-labeled oligoprobe, specific for somatostatin sst2 receptor mRNA. We observed a transient expression of SRIF receptors, labeled with [125I]LTT-SRIF-28, [125I]Tyr3-octreotide, and [125I]CGP 23996 (only in the presence of Mg2+ ions), in all principal auditory nuclei during neonatal development. In the adult rats, however, only the inferior colliculus displayed significant SRIF receptor binding. A very similar spatiotemporal labeling pattern was found for sst2 receptor mRNA. Our in situ hybridization data, together with those on ligand binding, suggest a predominantly transient expression of sst2 receptors in the auditory system. Since sst2 sites (and possibly sst3 and sst5) as well as SRIF itself appear to be co-expressed during a period when synapse maturation occurs, we suggest that sst2 receptors are involved in this process of the developing auditory system.

The messenger RNA (mRNA) expression of somatostatin (SRIF) receptors SSTR-1, SSTR-2, SSTR-3, SSTR-4 and SSTR-5 (called sst(1-5), now) was studied in rat brain between embryonic day 17 (E17) and post-natal day 5 (P5) by in situ hybridization histochemistry and compared to that of adult rats. sst(1) receptor mRNA expression was very low and restricted at E17, spread out at E18, to reach very high levels comparable to that of adult at P5 (e.g. in temia tecta, posteromedial cortical amygdaloid nucleus, subiculum). At E17/E18, sst(2) receptor mRNA expression was low and limited (telencephalon); significant levels were present at P5 in allocortex, hippocampus, locus coeruleus, similarly to adult brain. sst(3) receptor mRNA was high at E17 in most brain regions, and almost as ubiquitous as in adult brain at P5. sst(4) receptor mRNA was apparently absent at E17, with low levels in the hippocampus, amygdala and habenula at E18; a wider distribution, especially in the hippocampus and cerebral cortex was observed at P5, similar to that of adult. sst(5) receptor mRNA was not detected at E17 and negligible at E18; low levels were found in the cortex, hippocampus and cerebellum at P5. However, in adult brain, only the cerebellum and hind-brain showed some sst(5) receptor mRNA transcripts. The presence and distribution of SRIF receptor mRNAs differs substantially in embryo and adult brain. Some mRNAs are present throughout development, while others proceed only postnatally to the adult form. There are striking differences within and between the different SRIF receptor mRNAs, suggesting a role in neurogenesis for some SRIF receptors (e.g. sst(2)) However, mRNA and protein levels do not necessarily correlate.

The aim of the work was to investigate the effects of somatostatin analogs acting selectively on sst1 (BIM-23926), sst2 (BIM-23120) and sst5 (BIM-23206) receptor subtypes on the viability of “clinically non-functioning” pituitary adenomas in vitro. The effects of native SST (SST-14), a SST/DA chimera (BIM-23A387) and a D2-dopamine receptor agonist bromocriptine (BC) were also examined. The study was performed on 10 surgically removed pituitary macroadenomas, diagnosed before surgery as “non-functioning”. A part of each tumor was mechanically dispersed and digested with collagenase to isolate the tumoral cells. Another part of each tumor was fixed, embedded in paraffin and immunostained to reveal the pituitary hormones and SST receptor subtypes (sst1, sst2A, sst2B, sst3, sst4, sst5). The tumoral cell suspensions were incubated for 24 h with the substances mentioned above. The quantity of viable cells was estimated using the EZ4U system. The results were compared with the immunohistochemical evaluation of the hormonal profile of adenoma and the sst receptor subtype immunoreactivities present.The findings indicate that selective sst1, sst2 and sst5 receptors agonists, SST/DA chimera and D2-dopamine receptor agonist bromocriptine affect the viability of some, but not all, “clinically non-functioning” pituitary adenomas in vitro. The most effective was bromocriptine. The investigated somatostatin analogs including SST/DA chimera exerted roughly similar inhibitory effects. Further studies are needed to fully evaluate the potential usefulness of these compounds in the pharmacological treatment of “non-functioning” pituitary tumors.