3H-Spiroperidol (Spiperone) Binding Sites in Rat Adrenal Glomerulosa Cells

3H-spiperone, a dopaminergic antagonist, was used to study binding sites in rat adrenal glomerulosa membrane. The equilibrium dissociation constant (Kd) and binding capacity for 3H-spiperone binding were 2.2 nM and 268 fmol/mg protein, respectively. Determination of the Kd by kinetic studies provided a value of 2.6 nM, which corresponded closely to the Kd estimated by equilibrium studies. In a study of the subcellular distribution of dopamine receptors in adrenal glomerulosa cells, 3H-spiperone binding activity at the interface of density 1.14 to 1.16 accounted for 60% of the total activity in all fractions. These dopaminergic binding sites in adrenal glomerulosa cells may modulate aldosterone secretion induced by antidopaminergic agents

A Radioimmunoassay for Rat Serum Corticosterone

A reliable radioimmunoassay for rat serum corticosterone has been developed. 25 μ1 of diluted serum (1:100) was assayed with a specific antiserum raised against corticosterone-21-hemisuccinate conjugated to bovine serum albumin. The within-assay and between-assay coefficients of variation were 7.1% and 13.9%, respectively. The mean serum corticosterone concentration was 65.3±6.0 ng/ml (n=10). The corticosterone level increased to 208.4 ±23.7 ng/ml after ACTH administration, and was suppressed to the limit of assay sensitivity after dexamethasone administration

Steroidogenesis in Isolated Adrenal Cells of Rat

A reliable and reproducible system for the isolation of rat adrenal cells was developed, using 0.25% trypsin for cell dispersion. The suspending cell in Krebs-Ringer bicarbonate buffer containing 0.2% glucose and 0.5% bovine serum albumin was incubated for 120 minutes at 37℃ under 95% 02 and 5% C02. Corticosterone production induced by synthetic 1-24ACTH showed a dose-related increase in decapsular cells. The precision of the inter-experiment of corticosterone production was 5.0% (average coefficients of variation)

Analysis of Human Insulin Analogues in Vitro, Using Gel Chromatographic Method

The incubation medium and incubated human pancreas were gel chromatographed on the Bio-Gel P-30 column after extraction with acid ethanol. The extracted immunoreactive insulin (IRI) was successfully separated into two peaks at the position of 6000 molecular weight region. These two peaks corresponded to those which were detected in human serum. These findings suggest that the two groups of insulin are directly secreted from human pancreatic tissue. But the incorporated [3H] leucine peak into acid-ethanol extractable protein did not elute out at the same position as each insulin peak. Therefore, the measurement of [3H] leucine incorporation into acid-ethanol extractable protein is not a good indicator to evaluate insulin biosynthesis

Distribution and Elimination of Insulin and C-peptide in a Benign Insulinoma Patient

The distribution and elimination of insulin and C-peptide was evaluated in a case of benign insulinoma, using the method of gel chromatography. The significant differences between total Immunoreactive insulin (IRI) level and the level of Peak I plus Peak II of IRI was noticed in splenic vein. This fact suggested that intermediate and/or abnormal IRI could be released from the tumor. In order to diagnose a hypoglycemic patient with completely normal IRI and CPR level in peripheral blood, the gel chromatographic separation of IRI from splenic and/or portal blood could be useful