Biomedical subjects
L J Miller
Publications and source records attributed to L J Miller.
Complex role of protein kinase C in mediating the supramaximal inhibition of pancreatic secretion observed with cholecystokinin.
Protein kinase C appears to play an important, yet complex role in the supramaximal inhibition of pancreatic acinar cell secretion observed in response to cholecystokinin (CCK). The addition of protein kinase C activation to the concentration-response curve of a partial agonist acting at the CCK receptor (a phenethyl ester analogue of CCK), transforms a curve without supramaximal inhibition to a full agonist curve typical of CCK. This effect can be elicited by low concentrations of phorbol ester (50pM to 1nM 12-0-tetradecanoyl-phorbol-13-acetate) or by hormonal agonists (0.1 microM carbamylcholine, 10pM bombesin, 1pM CCK-8) which activate protein kinase C, but not by agonists acting via alternate second messengers (VIP). Of interest, this effect is dependent on preincubation of the acinar cells with the protein kinase C activator at 37 degrees C, with the effect rapidly reversed by transient exposure of the cells to lower temperature. This is consistent with mediation by a phosphorylation event. However, the requirement for an extended (greater than 15 min) preincubation period when using minimal kinase activation suggests that this phenomenon is more complicated than a simple bimolecular phosphorylation event and likely includes a series of events such as translocation of substrates and/or enzymes involved.
Expression cloning and characterization of the canine parietal cell gastrin receptor.
Gastrin is an important stimulant of acid secretion by gastric parietal cells and is structurally related to the peptide hormone cholecystokinin (CCK). The pharmacologic properties of the parietal cell gastrin receptor are very similar to the predominant CCK receptor in the brain, CCK-B. Neither the gastrin nor the CCK-B receptor have been cloned thus far, making it difficult to resolve whether these two receptors are distinct. We have isolated a clone encoding the canine gastrin receptor by screening a parietal cell cDNA expression library using a radioligand-binding strategy. Nucleotide sequence analysis revealed an open reading frame encoding a 453-amino acid protein with seven putative hydrophobic transmembrane domains and significant homology with members of the beta-adrenergic family of G protein-coupled receptors. The expressed recombinant receptor shows the same binding specificity for gastrin/CCK agonists and antagonists as the canine parietal cell receptor. Gastrin-stimulated phosphatidylinositol hydrolysis and intracellular Ca2+ mobilization in COS-7 cells expressing the cloned receptor suggest second-messenger signaling through phospholipase C. Affinity labeling of the expressed receptor in COS-7 cells revealed a protein identical in size to the native parietal cell receptor. Gastrin receptor transcripts were identified by high-stringency RNA blot analysis in both parietal cells and cerebral cortex, suggesting that the gastrin and CCK-B receptors are either highly homologous or identical.
Binding of a phenethyl ester analogue of cholecystokinin to the solubilized pancreatic cholecystokinin receptor: use in ligand-affinity chromatography.
Pancreatic acinar cells have both high and low affinity receptors for cholecystokinin (CCK), yet their membranes appear to possess only a single class of binding sites. Recently, gallbladder membrane CCK receptors were shown to undergo inter-conversion between two affinity states dependent on G protein coupling. Keys for that observation were the differential binding affinities of CCK and a phenethyl ester analogue of CCK (OPE), with the high affinity state binding CCK with higher affinity than OPE, and the low affinity state binding OPE with higher affinity than CCK. Here, we performed analogous experiments using these ligands and both pancreatic membranes and a solubilized preparation. Both preparations were found to have only single affinity states of this receptor. However, the state on membranes had a higher affinity for CCK than for OPE, and that on the solubilized preparation had a higher affinity for OPE than for CCK. This supports the hypothesis that the ternary complex of ligand-receptor-G protein found in membranes represents the high affinity state of this receptor, while the uncoupled form of this receptor after solubilization represents its low affinity state. The high affinity of OPE for the solubilized receptor can be utilized in a purification strategy to follow receptor-bearing fractions and to provide an efficient and specific affinity-binding step.
A historical perspective of gastrointestinal endocrinology--the new age of molecular receptorology.
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The Zollinger-Ellison syndrome. A collective surgical experience.
A retrospective study of 90 surgically treated patients with the Zollinger-Ellison syndrome seen from 1958 through 1990 was performed. Fifteen patients had Zollinger-Ellison syndrome as a manifestation of multiple endocrine neoplasia type I. Preoperative tumor localization was positive in 46% of 54 patients studied. Gastrinomas were identified in 66% of patients, 38% of the tumors being malignant. Postoperative eugastrinemia was achieved in 11% of patients after a variety of surgical procedures. Exploratory laparotomy provides the only chance for cure and identifies the significant prognostic factors associated with long-term patient survival: small tumor size, extrapancreatic primary, and absence of tumor metastases.
Situational influence on development of delusions of pregnancy in a man.
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Ifosfamide-induced neurotoxicity: a case report and review of the literature.
OBJECTIVE: To review published abstracts, journal articles, and case reports for evidence of ifosfamide-induced neurotoxicity in both adult and pediatric populations. DATA SOURCES: Peer-reviewed journal articles (October 1985 through August 1990) obtained through a computer literature search, with subsequent bibliography scanning. STUDY SELECTION: We identified 34 reports that specifically addressed neurotoxicity related to ifosfamide therapy. By consensus of the authors, 8 of these reports were excluded due to a small study population, duplication of age of data. DATA EXTRACTION: Studies were assessed by the first author and verified by the second author. Several colleagues also contributed to the analysis. DATA ANALYSIS: Risk factors and mechanisms behind this potentially fatal neurotoxic reaction remain speculative and, in some cases, contradictory. The predictive capability of a published nomogram is restricted by differences in dosage regimens and encephalopathic classifications. Currently, the best prevention against neurotoxicity is little or no use of concurrent medications that have central nervous system effects. CONCLUSIONS: Further studies should address the influence of other nephrotoxic agents (e.g., cisplatin, aminoglycosides) on the incidence of ifosfamide-induced neurotoxicity.
The role of the histotechnician in Mohs surgery.
Critical elements of Mohs micrographic surgery are performed by the histotechnician in the frozen section laboratory. This article provides an overview of the essential role of the histotechnician as a member of the micrographic surgical team.
Agonist-regulated phosphorylation of the pancreatic cholecystokinin receptor.
The present study was undertaken to determine if the cholecystokinin (CCK) receptor may be phosphorylated, and to gain insight into its regulation. For this, the ATP pool of rat pancreatic acini was prelabeled with 32P, and the cells were stimulated with various secretagogues. CCK receptors from treated cells were enriched by sequential fractionation to produce plasmalemma, and subsequent solubilization and lectin-affinity chromatography. This protocol detected a phosphorylated Mr = 85,000-95,000 plasma membrane glycoprotein with features similar to the CCK receptor. Phosphorylation of this protein occurred rapidly (less than 2 min) and in a concentration-dependent manner in response to CCK, and was inhibited by the CCK receptor antagonist L-364,718. Further evidence that this represented the CCK receptor included comigration of phosphorylated and CCK radioligand affinity-labeled proteins on sodium dodecyl sulfate-polyacrylamide gels, both in native forms and after endoglycosidase F deglycosylation, and the specific adsorption of the phosphoprotein to a CCK analogue affinity resin. Phosphorylation occurred predominantly on serine residues of the receptor protein. Phosphorylation of this protein was also enhanced in response to other secretagogues which, like CCK, stimulate a cascade leading to protein kinase C activation, and in response to direct activation of this enzyme by 12-O-tetradecanoylphorbol 13-acetate. Thus, the pancreatic CCK receptor is phosphorylated in a regulated manner, in response to both homologous and heterologous secretagogues, and to protein kinase C activation.
Use of photoaffinity probes containing poly(ethylene glycol) spacers for topographical mapping of the cholecystokinin receptor complex.
To further define the structure of the pancreatic cholecystokinin (CCK) receptor and the topographical distance relationships between its subunits, we developed a series of monofunctional photoaffinity probes in which a fixed receptor-binding domain was separated from a photolabile nitrophenylacetamido group by defined lengths of a flexible spacer. The well-characterized CCK receptor radioligand 125I-D-Tyr-Gly-[(Nle28,31)CCK-26-33] provided the receptor-binding component of the probes, while the polymer poly(ethylene glycol) (2, 4, 7, and 10 monomer units long) was used as the spacer. The patterns of affinity labeling of rat pancreatic plasma membranes were examined as a function of spacer length. This ranged from 7.3 to 16.2 A, as calculated by root-mean-square end-to-end distances and validated experimentally by time-resolved fluorescence resonance energy transfer measurements. All probes in the series specifically labeled the Mr = 85,000-95,000 glycoprotein with Mr = 42,000 core, which has been proposed to contain the hormone recognition site. In addition, when the spacer length reached 16.2 A, membrane proteins of Mr = 80,000 and Mr = 40,000 were specifically labeled. The product of endo-beta-N-acetylglucosaminidase F digestion of the Mr = 80,000 protein was Mr = 65,000, similar to a protein previously identified in affinity labeling experiments using a CCK-33-based probe. These observations are consistent with the Mr = 85,000-95,000 pancreatic protein representing the hormone-binding subunit of the CCK receptor, while proteins of Mr = 80,000 and Mr = 40,000 may represent noncovalently associated subunits sited within 16.2 A of the binding domain.(ABSTRACT TRUNCATED AT 250 WORDS)
Enteric neuronal autoantibodies in pseudoobstruction with small-cell lung carcinoma.
Severe gastrointestinal dysmotility is a newly recognized paraneoplastic syndrome that occurs with small-cell lung carcinoma. Thirty-four patients with small-cell carcinoma, of whom 5 had chronic intestinal pseudoobstruction and 29 had no digestive symptoms, were studied serologically. Four of the 5 patients with gut dysmotility had immunoglobulin G antibodies reactive with neurons of the myenteric and submucosal plexuses of jejunum and stomach in an indirect immunofluorescence assay. Antibodies of this type were not found in any of the 29 patients who had no gut dysmotility, nor were they found in patients with chronic idiopathic intestinal pseudoobstruction (n = 8), ovarian cancer (n = 20), or epilepsy (n = 4) or in normal subjects (n = 9). In 4 of the patients with paraneoplastic pseudoobstruction, antibodies in highly diluted serum (1:4000-1:8000) bound selectively to nuclei and cytoplasm of neuronal elements in the gut. This novel autoantibody activity suggests that intestinal pseudoobstruction occurring in patients with small-cell carcinoma may have an autoimmune basis. From a clinical standpoint, serological testing offers a simple means for determining which patients with gut dysmotility syndromes may have associated small-cell carcinoma, thereby enabling earlier diagnosis and treatment of the tumor.
Abnormal gallbladder motility in irritable bowel syndrome: evidence for target-organ defect.
We have described previously that the gallbladder responds abnormally to infusions of cholecystokinin octapeptide (CCK-8) in patients with irritable bowel syndrome (IBS). To confirm these results and to examine the possible mechanisms, patients with IBS and predominant symptoms of diarrhea or constipation were compared with matched controls. During infusions of CCK-8 at one of three doses, the response of the gallbladder was measured ultrasonographically. The levels of CCK-8 reached in the peripheral circulation and degradation of the peptide in vitro and in vivo were used to evaluate metabolism of cholecystokinin. We confirmed that the gallbladders of patients with IBS responded abnormally to CCK-8; however, the differences were not due to any prereceptor event. Instead, this abnormality in IBS must be explained by an atypical response at the level of the target tissues.
Impact of insurance review process on inpatient care.
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Risks of restraints versus psychotropic medication for pregnant patients.
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Maladaptive denial of pregnancy.
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Clinical strategies for the use of psychotropic drugs during pregnancy.
The treatment of major psychiatric disorders during pregnancy is becoming an increasing challenge for the clinician as fertility rates rise among mentally ill women. When prescribing psychotropic medication, the risk/benefit analysis comes to include the risks of the medication to the fetus, special risks to the mother because of the pregnant state, and the risks to both mother and fetus of the untreated psychiatric disorder. This article will review what is known about the risks to mother and fetus of antipsychotic, antidepressant, anxiolytic, mood stabilizing and antiparkinsonian agents, and will present treatment strategies designed to minimize those risks.
The gall bladder cholecystokinin receptor exists in two guanine nucleotide-binding protein-regulated affinity states.
To study proximal events in cholecystokinin (CCK) action on bovine gall bladder smooth muscle, we used the hormone analogue D-Tyr-Gly-[(N1e28,31)CCK-26-32]-phenethyl ester (OPE), which has unique biological properties. This fully efficacious agonist differs from native CCK by not expressing supramaximal inhibition of cell shortening, yet it clearly interacts with the same receptor molecule. This was demonstrated in binding and affinity labeling studies, where both peptides label the same Mr 70,000-85,000 protein and both fully compete for binding of the other ligand. Further, its relatively high affinity for the low affinity CCK receptor permits the clear demonstration of two affinity states of a CCK receptor on a membrane preparation and makes possible evaluation of the molecular basis of these affinity states and their regulation. Analysis of homologous and heterologous binding curves performed with both CCK and OPE peptides and radioligands demonstrated the presence of two affinity states, with CCK being able to distinguish them (Kd1 = 0.48 +/- 0.04 nM and Kd2 = 56.5 +/- 7.4 nM) and OPE recognizing them equally (Kd1 = 0.94 +/- 0.31 nM and Kd2 = 0.96 +/- 0.23 nM). In the presence of nonhydrolyzable GTP analogues, there was a shift in distribution of receptors toward the low affinity state, with the total number of receptors and their absolute affinities for each peptide remaining constant. Thus, the gall bladder CCK receptor is a single molecule capable of assuming two interconvertible affinity states, regulated by a guanine nucleotide-binding protein. Two full agonists are capable of interacting with this molecule to yield different biological responses via different molecular events.