The stability and metabolism of intravenously administered neurotensin in the rat

The clearance and metabolism of synthetic and tritiated (³H) neurotensin (NT) were studied following its intravenous injection in a pharmacologic dose (500 pmol/kg) into anesthesized rats. Immunoreactive NT (iNT), measured in a radioimmunoassay (RIA) with use of a carboxyl-(C)-terminal directed antiserum, displayed an apparent half-life ($\text{t}\text{1}\text{2}$) of 0.55 min, while that measured by an amino-(N)-terminal directed antiserum had a $\text{t}\text{1}\text{2}$ of 5 min. The radiolabel from injected ³H-NT (³H on Tyr^3,11) had a $\text{t}\text{1}\text{2}$ of 6.5 min. High-pressure liquid chromatography of extracts of plasma obtained from the circulation 0.5–3 min after injection of NT and ³H-NT showed the presence of NT and the generation mainly of the fragments NT^1–8, NT^1–11, and NT^9–13, as well as free ³H-labeled tyrosine. The apparent half-lives of intravenously injected synthetic NT^1–8, NT^1–11 and NT^1–12 measured with the N-terminal RIA were 9, 5 and 5 min, respectively, while that for NT^9–13 was less than 0.5 min. These results indicate that exogenously injected NT is rapidly metabolized to form N-terminal fragments which are cleared more slowly than NT. These findings suggest that use of N-terminal antisera to detect the release of endogenous NT into the circulation is likely to yield measurements of the fragments NT^1–8 and NT^1–11 which thus far have been found to be biologically inactive.     

Ultrastructure of the gut neurotensin cell

Summary In mammals, neurotensin cells occur scattered in the epithelium of the jejunum-ileum. In chicken, neurotensin cells are abundant in the region of the gizzard-duodenal junction (antrum) where they occur intermingled with numerous somatostatin and gastrin cells. The neurotensin cells in chicken, dog and man were identified at the electron microscopic level by immunocytochemistry, using the consecutive semithin/ultrathin section technique. They contain numerous electron dense cytoplasmic granules, predominantly in the basal portion of the cell. It was shown that these granules are the storage site for neurotensin. The neurotensin granules are round, highly electron dense and of about the same size in the different species examined (mean diameter 260–290 nm). in dog and man the granules have a tightly applied surrounding membrane while in the chicken a relatively electron lucent zone separates the electron dense core from the granule membrane. The ultrastructure of the neurotensin granules in chicken is some-what reminiscent of that of the gastrin granules. The mean diameter of the gastrin granules in chicken antrum is 230 nm; for the somatostatin granules the mean diameter is 305 nm.     

Ultrastructure of the gut neurotensin cell.

In mammals, neurotensin cells occur scattered in the epithelium of the jejunum-ileum. In chicken, neurotensin cells are abundant in the region of the gizzard-duodenal junction (antrum) where they occur intermingled with numerous somatostatin and gastrin cells. The neurotensin cells in chicken, dog and man were identified at the electron microscopic level by immunocytochemistry, using the consecutive semithin/ultrathin section technique. They contain numerous electron dense cytoplasmic granules, pre-dominantly in the basal portion of the cell. It was shown that these granules are the storage site for neurotensin. The neurotensin granules are round, highly electron dense and of about the same size in the different species examined (mean diameter 260--290 nm). In dog and man the granules have a tightly applied surrounding membrane while in the chicken a relatively electron lucent zone separates the electron dense core from the granule membrane. The ultrastructure of the neurotensin granules in chicken is somewhat reminiscent of that of the gastrin granules. The mean diameter of the gastrin granules in chicken antrum is 230 nm; for the somatostatin granules the mean diameter is 305 nm.     

Isolation and identification of a polypeptide in the Hsp 70 family that binds substance P.

During the course of an attempt to purify the substance P (SP) receptor from horse salivary glands by substance P-affinity chromatography, a polypeptide of Mr = 78,000 was isolated. The first fifteen amino acid residues at the amino terminus were determined and, unexpectedly, were found to be identical with the amino terminus of a glucose-regulated protein (GRP) of the same molecular weight, a protein that has been identified as a member of the heat shock protein family. This finding raises the intriguing possibility that SP may interact in vivo with GRPs and other members of the heat shock protein family and play a role in modulating their biological activities.     

The fluorescence and bright field microscopic demonstration of cathepsin B in human fibroblasts

Cathepsin B was demonstrated cytochemically in human fibroblasts with Z-Ala-Arg-Arg-2-(4-methoxy)naphtylamide as substrate. The enzyme was visualized in the bright field microscope with the diazonium salt Fast Blue B as coupling reagent and in the fluorescence microscope with 5-nitrosalicylaldehyde. With both methods cathepsin B was found in small granules distributed throughout the cytoplasm.     

1.5 MHz ultrasound irradiation of human lymphocytes.

Human lymphocyte cultures are exposed to 1.5 MHz continuous wave ultrasound, and it is demonstrated that cell death, as monitored by pyknosis, follows immediately on sonication at intensities within the usual therapeutic range (less than 1.7 W/cm2,spatial average). The number of cells affected is determined by the ultrasound intensity only, but the rate at which they proceed through their pyknosis cycle is modified by both the intensity and the duration of exposure. There is a clear indication of an intensity threshold for the effect approximately 1.1 W/cm2. Pulsed 1.5 MHz ultrasound (70 mus, 1 :1 pulses, 1.7 W/cm2 space-time average) results in a 15-20 hour delay in the measurable response to sonication. It is shown that the intracellular presence of a lysosomal-enzyme inhibitor strongly modifies the course of the ultrasound action. Evidence is presented to suggest that the basic interaction mechanism is via a cavitation process, but there are some difficulties with this interpretation, which are also discussed.     

Characterization of the Substance P (NK-1) Receptor in Tunicamycin-Treated Transfected Cells Using a Photoaffinity Analogue of Substance P

Abstract: Chinese hamster ovary cells expressing the N -glycosylated substance P (NK-1) receptor were treated with the glycosylation inhibitor tunicamycin and photolabeled with ¹²⁵I-Bolton-Hunter- p -benzoyl- l -phenylalanine⁸-substance P. Two radioactive proteins of M_r 80,000 and 46,000, representing the glycosylated and nonglycosylated substance P (NK-1) receptor, respectively, were observed. The IC₅₀ for the inhibition of photolabeling of both receptor forms was 0.3 ± 0.1 n M for substance P and 30 ± 5 n M for neurokinin A (substance K). Thus, glycosylation of the substance P (NK-1) receptor has no detectable effect on the affinity of the substance P (NK-1) receptor for substance P or neurokinin A (substance K).