Development of antibodies against the rat brain somatostatin receptor.

Théveniau, Magali; Rens-Domiano, Stephanie; Law, Susan F.; Rougon, Geneviève; Reisine, Terry

doi:10.1073/pnas.89.10.4314

Cited by 16 publications

(11 citation statements)

References 18 publications

(37 reference statements)

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“…The lack of mRNA encoding the different cloned SRIF receptors and the low levels or absence of SSTR2 immunoreactivity in the striatum suggests that this brain region may express a novel SRIF receptor subtype that has not been cloned . Consistent with this hypothesis, we (Theveniau et al ., 1992) generated polyclonal antiserum against partially purified rat brain SRIF receptors that strongly reacted with a rat striatal SRIF receptor. This polyclonal antiserum does not cross-react with any of the cloned SRIF receptors (M. A .…”

Section: Discussionsupporting

confidence: 85%

“…Theveniau and T. Reisine, unpublished results), indicating that it recognizes a novel SRIF receptor subtype . The further identification of the rat striatal SRIF receptor should be clarified after its cloning, possibly through the use of our recently developed polyclonal antiserum (Theveniau et al, 1992) .…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, affinity purification of rat brain SRIF receptors has revealed a protein of 60 kDa . In addition, antibodies (F4) generated against partially purified rat brain SRIF receptors only react with 60-kDa proteins from rat brain (Theveniau et al, 1992) . These lower molecular mass SRIF receptors may correspond to a subtype different from SSTR2 .…”

Section: Discussionmentioning

confidence: 99%

“…For immunoprecipitation of rat brain SSTR2, a gel-filtration stage was added before immunoprecipitation as described by Law et al . (1991) and Theveniau et al . (1992) .…”

Section: Peptide-directed Antibodies Detected Sstr2 Isoformsmentioning

confidence: 97%

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Immunological Detection of Isoforms of the Somatostatin Receptor Subtype, SSTR2

Théveniau

Yasuda

Bell

et al. 1994

Journal of Neurochemistry

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Somatostatin (SRIF) induces its diverse physiological actions through interactions with different receptor subtypes. Multiple SRIF receptor subtypes have recently been cloned. To analyze the physical properties of receptor subtype SSTR2, two different peptide‐directed antibodies were generated against SSTR2. Antibody “2e3,” directed against the peptide SSCTINWPGESGAWYT (residues 191–206), corresponding to a region in the predicted third extracellular domain of mouse SSTR2, and antibody “2i4,” directed against the peptide SGTEDGERSDS (residues 333–343) from the predicted cytoplasmic tail of mouse SSTR2, were developed. In Chinese hamster ovary (CHO) cells stably expressing the mouse SSTR2 gene (CHOB), the antibody 2e3 recognized specifically a protein of 93‐kDa protein by immunoblotting. No specific immunoreactivity was detected by 2e3 in nontransfected CHO cells or CHO cells stably expressing vector alone or human SSTR1 or mouse SSTR3 genes. The antibody 2i4 specifically immunoprecipitated SSTR2 solubilized from CHOB cells that could be labeled with the SSTR2‐specific ligand 125I‐MK‐678. Furthermore, both 2e3 and 2i4 specifically immunoprecipitated 93‐kDa [35S]methionine‐labeled proteins from CHOB cells, indicating that they recognize the same proteins. In contrast to studies in CHOB cells, immunoblotting studies showed that 2e3 detected specifically a single 148‐kDa protein from different regions of the rat brain that have previously been shown to express high levels of SSTR2 mRNA and SRIF receptors with high affinity for 125I‐MK‐678. In contrast, no immunoreactivity was detected in rat kidney, liver, or lung, which do not express SSTR2. No 93‐kDa protein was detected specifically in the rat brain. The 148‐kDa protein detected by 2e3 is an SRIF receptor because 2e3 and 2i4 specifically immunoprecipitated solubilized rat brain SRIF receptors that could be reversibly labeled with 125I‐MK‐678. As in rat brain, 2e3 interacted specifically with a single 148‐kDa protein in rat pituitary, in the rat pancreatic cell line AR42J, and in the HEK 293 cell line derived from human kidney, all of which express SSTR2 mRNA and SRIF receptors with high affinity for 125I‐MK‐678. These findings indicate that rat brain and pituitary, as well as a pancreatic and a kidney cell line, express primarily a form of SSTR2 different from CHOB cells. The multiple forms of SSTR2 may result from differential post‐translational processing of SSTR2 because 2e3 immunoprecipitated 41‐kDa in vitro translation products generated from mRNA extracted from CHOB and AR42J cells. This 41‐kDa protein has the predicted size of unprocessed SSTR2. These results demonstrate that 2e3 and 2i4 antibodies interact specifically with SSTR2. Detection of two different size proteins by the SSTR2 peptide‐directed antibodies suggests the existence of multiple forms of SSTR2.

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Section: Discussionsupporting

confidence: 85%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

“…For immunoprecipitation of rat brain SSTR2, a gel-filtration stage was added before immunoprecipitation as described by Law et al . (1991) and Theveniau et al . (1992) .…”

Section: Peptide-directed Antibodies Detected Sstr2 Isoformsmentioning

confidence: 97%

See 2 more Smart Citations

Immunological Detection of Isoforms of the Somatostatin Receptor Subtype, SSTR2

Théveniau

Yasuda

Bell

et al. 1994

Journal of Neurochemistry

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“…Previous studies by Gonzalez et al (1989) reported transient expression of ]SRIF binding sites early in cerebellar development. In the present study, we have investigated the developmental pattern of expression of SRIF receptors in the cerebellum and other rat brain regions by using our recently developed antibody against a 60-kDa rat brain SRIF receptor to detect the receptor by immunoblotting (Theveniau et al, 1992). This approach allows us to determine whether qualitative changes occur in receptor level as well as whether physical changes occur in the receptor during development.…”

mentioning

confidence: 99%

Developmental Changes in Expression of a 60‐kDa Somatostatin Receptor Immunoreactivity in the Rat Brain

Théveniau

Reisine

1993

Journal of Neurochemistry

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The neuropeptide somatostatin (SRIF) exerts several important physiological actions in the adult CNS through interactions with membrane-bound receptors. SRIF expression is developmentally regulated and this regulation is most apparent in the cerebellum, where SRIF immunoreactivity is expressed at early postnatal ages and then disappears toward adulthood. The transitory nature of SRIF expression at a time of major changes in cerebellum suggests that this peptide may have a role in cerebellar development. To further investigate the role of the SRIF transmitter system during development, we have examined the levels of expression of SRIF receptors in the developing rat brain by immunoblotting using antiserum selective for a 60-kDa brain SRIF receptor. In whole rat brain, SRIF receptor immunoreactivity first appears at embryonic day 13 (E13), is elevated at E16, increases at birth, peaks at early postnatal ages, and then gradually declines with age. No apparent changes in size of the receptor occur with age. No consistent changes in levels of SRIF receptor immunoreactivity are detected from early postnatal ages to adulthood in the hippocampus, cerebral cortex, and striatum, but levels gradually decline in the hypothalamus. In contrast, SRIF receptor immunoreactivity is expressed transiently in cerebellum. SRIF receptor immunoreactivity is detectable in cerebellum at E16, increases in levels at birth, is apparent from postnatal day 3 to postnatal day 8, and then disappears. The transitory nature of SRIF receptor expression in cerebellum is unique and parallels the expression of SRIF immunoreactivity in this brain region. These findings support the hypothesis that SRIF has a role in cerebellar development.

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Heterogeneity in hepatic transport of somatostatin analog octapeptides

et al. 1994

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Hepatic transport of the synthetic somatostatin analog octreotide-SMS 201-995, (D)Phe-Cis-Phe-(D)Trp-Lys-Thr-Cis-Throl-and its novel derivative N-a-(a-D-glucosy1( 1-4)-1-deoxy-D-fructosyl)-octreotide -SDZ CO-611, ~-a-(a-D-g~ucosyl( 1-4)-l-deoxy-Dfructosyl)-(D)Phe-C&-Phe-(D)Trp-Lys-Thr-Cis-Throlwas studied. In rats SMS 201-995 showed a plasma elimination half-life of 1.2 & 0.2 hr; that of SDZ CO-611 was 1.9 +-0.3 hours. Within 120 min 66% of a mesenterically injected 4.4-nmol dose of SMS 201-995 was excreted in bile, but only 5.3% of SDZ CO-611 was excreted in bile. Biliary concentration of SMS 201-995 showed a maximum enrichment of 540-fold & 75-fold over peripheral blood concentration, indicating hepatic transport mechanisms different from simple diffusion. Comparison of plasma profiles of both peptides after mesenteric and femoral administration demonstrated the relative importance of hepatic extraction for SMS 201-995 but not for SDZ CO-611. The mode of extraction was studied by means of multiple-indicator dilution in isolated perfused rat liver, with inulin as nonpermeable marker. Ratio plots, ln([inulin]/[peptide]) vs. time, exhibited decreasing slopes for SMS 201-995, suggesting very rapid binding to hepatocyte membranes. The slope of the ratio plot of (inulin/SDZ CO-611) was almost zero even at low doses (down to 0.2 Fg), implying mainly extracellular distribution and nonhepatic elimination. Binding assays indicated the absence of somatostatin receptors in sinusoidal hepatocyte membranes. However, SMS 201-995 and SDZ CO-611 bound with high affinity to somatostatin receptors in rat cortical membranes. Multiple-indicator dilution experiments in presence of increasing cholyltaurine concentrations suggested an interaction of SMS 201-995 with sinusoidal bile salt transport. In isolated hepatocytes, uptake of SMS 201-995 was satu-

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Development of antibodies against the rat brain somatostatin receptor.

Cited by 16 publications

References 18 publications

Immunological Detection of Isoforms of the Somatostatin Receptor Subtype, SSTR2

Immunological Detection of Isoforms of the Somatostatin Receptor Subtype, SSTR2

Developmental Changes in Expression of a 60‐kDa Somatostatin Receptor Immunoreactivity in the Rat Brain

Heterogeneity in hepatic transport of somatostatin analog octapeptides

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