Murray Saffran

Insulin and the gastrointestinal tract

Abstract To provide a less cumbersome and more socially accepted form of insulin treatment than subcutaneous injections, we have designed an azopolymer system to deliver insulin with an absorption enhancer to the upper colon. In pancreatectomized dogs repeated oral doses of insulin in azopolymer-coated capsules lower the diabetic hyperglycemia to near normal values. Direct visualization of capsules containing radionuclides and coated with two new batches of azopolymer demonstrated that the capsules either passed intact through the gut or were opened in the small intestine. Direct visualization and insulin delivery using the same azopolymer will be necessary to locate the site of insulin delivery, but there remains the possibility that the insulin was delivered within the small intestine. To understand the effect of large doses of insulin delivered to the upper GI tract, an insulin solution was substituted for the drinking water of normal and diabetic rats. This produced temporary decreases in blood glucose levels, showing that in rats some absorption of insulin takes place above the colon. However, these rats became hyperphagic and lost weight. On post mortem examination the gut was distended with undigested food. Insulin seemed to inhibit processing of the food by the gut, in confirmation of observations by Elliasson and coworkers in human volunteers. The inhibitory effect of insulin on the gut, coupled with the presence of insulin receptors on the mucosal side of the gut and the presence of other pancreatic peptides in the gut, suggested to us that the gut may be able to make insulin. Accordingly, we looked for and found immunocytochemical evidence for preformed insulin in crypt cells in the colon and stomach, as well as the mRNAs for both rat insulins in similar cells. Gut insulin may be involved in the response of the gastrointestinal tract to food. Caution must be exerted in the introduction of high concentrations of insulin into the gastrointestinal tract. The same may be true of other powerful natural agents for which an oral delivery system would be desirable.

A Semi-Automated in vitro Assay for CRF: Activities of Peptides Related to Oxytocin and Vasopressin

The CRF-like activity of peptides related to vasopressin and oxytocin was estimated using a semi-automated in vitro bioassay. Rat pituitary halves were incubated for 1 h with the test material in 1.0 ml of Krebs-Ringer bicarbonate buffer. The ACTH released from the pituitary tissue was estimated using adrenal quarters in an automated superfusion system. The pituitary tissue was standardized with median eminence extract, and adrenal tissue was standardizeed with ACTH. Vasopressin and its analogues release ACTH in order of their pressor potency. Most of the ring or tail fragments of oxytocin and vasopressin did not stimulate the release of ACTH. In some experiments, deaminopressinamide and pressinoic acid had CRF-like activity.

Analysis of the Response to ACTH by Rat Adrenal in a Flowing System

Publisher Summary This chapter describes a flowing system for the study of adrenocortical function using rat tissue in vitro, in which the fluorometric measure of corticosterone is completely automated. In the study, the apparatus was assembled from standard pumps, tubing, and fittings supplied by Technicon Corporation and from common laboratory parts. The apparatus was calibrated by the passage of standard amounts of corticosterone. When adrenal tissue was introduced into the flowing system, there was an immediate high increase in fluorescence values. The flowing system was uniquely suited to studying the influence of the rate of dose administration of adrenocorticotropic hormone on the adrenal response. When a standard solution of corticosterone was introduced into the flowing system, there was no apparent response for approximately 20 min. The delays encountered when the source of corticosterone is adrenocortical tissue are the same as those with the standard solution of corticosterone.

Evidence for biosynthesis of preproinsulin in gut of rat

Glucagon and other pancreatic peptides are made in the gut, but there is little evidence for the formation of insulin. The demonstration of insulin receptors on the mucosa of gut epithelium suggests that there may be an autocrine or paracrine role for insulin made in the gut. Such insulin may control cell division, the secretion of other peptides from the same or neighboring cells, or motility and absorption. To search for the ability of the gut to make insulin, sections of freshly excised segments of rat gut were treated with an antiserum against porcine insulin. Intracellular immunore-activity appeared in glandular cells in the stomach and colon but not in the small intestine. Preproinsulin mRNA was detected in similar cells in the stomach and colon by in situ hybridization, using specific oligonucleotide probes. Rat preproinsulin 1 and 2 mRNAs were transcribed by reverse transcriptase to the corresponding cDNAs, which were then amplified by polymerase chain reaction, utilizing specific oligonucleotide primers. Restriction analysis confirmed the identity of rat preproinsulin 1 and 2 mRNA in the colon and rat preproinsulin 1 mRNA in the stomach. Neither was found in the small intestine. Base sequences of the cDNAs were identical to the coding regions of pancreatic rat preproinsulin 1 and 2 messages. These observations are strong evidence for the synthesis of preproinsulin in the gut of the rat.

Pressinoic acid: A peptide with potent corticotrophin-releasing activity

Abstract Pressinoic acid, a synthetic hexapeptide that corresponds to the ring of vasopressin, exhibited corticotrophin-releasing activity in , vitro in doses of 3 and 30 nanograms per ml. The related peptide, deaminopressinamide, released lesser amounts of corticotrophin in doses of 30 to 30,000 ng. Two other hexapeptides, pressinamide and deaminopressinoic acid, were devoid of corticotrophin-releasing activity at doses of 30 to 30,000 ng.

CORTICOTROPIN‐RELEASING FACTOR: ISOLATION AND CHEMICAL PROPERTIES*

The concept of corticotropin-releasing factor (CRF) and its control of the release of ACTH is enshrined in American literature as indicated by the above quotation from Kurt Vonnegut, Jr.k Breakfast of Champions. Our entry into the field of research relating to the control of the release of ACTH was inspired by our then chief, Dr. R. A. Cleghorn, with whom we worked at the Allan Memorial Institute of Psychiatry, McGill University in Montreal. Dr. Cleghorn and his group were interested in the relationship between mental*disease and endocrine function.2 He introduced us to the concept that the interface between the central nervous system and the endocrine system was located in the hypothalamus. The elegant summary of the knowledge in this area by G. W. Harris? provided us with the theoretical basis for our research. There seemed to be a connection between the stresses of life and mental status and the activation of the pituitary-adrenocortical system. According to the hypothesis (FIGURE 1 ), the hypothalamus responded to stress by secreting a chemical factor that was responsible for the release of ACTH from the anterior pituitary. Accordingly, we undertook the search for the hypothetical chemical agent. We reasoned that if the scheme in FIGURE 1 is correct, then the isolated pituitary gland should respond to the application of the hypothetical hypothalamic agent by releasing more ACTH. We therefore designed a test system

Oral Administration of Insulin: Imitating the Natural Pathway

Before the discovery of insulin by Banting and Best in 1921, a diagnosis of diabetes, especially in a young child, was a sentence of death. Since then, insulin has given the patient with diabetes mellitus many years of life at the expense of daily injections. So far no practical alternative has been developed to the injection route for administration of insulin. As time passed and the initial gratitude for the life-preserving effect of insulin became commonplace, the drawbacks of insulin injections became apparent. First of all there is the discomfort of the injection, now lessened by the development of thinner needles and needle lubricants, the danger of infection, mitigated by convenient alcohol swabs and the disposable needle and syringe, the desire for privacy, decreased somewhat by the development of injection pens and external pumps, and the bother of carrying around and caring for supplies of insulin solutions. With prolonged life came the dilemma of running out of suitable injection sites, as old ones became pin cushioned and less receptive to the needle. But more important than all of these drawbacks of injected insulin, the insulin is delivered to the wrong place.

Is insulin a factor in the genesis of the vascular complications of diabetes

Several recent studies report that insulin may be a pathogenic factor in cardiovascular disease and the vascular complications of diabetes. The route of administration of insulin may be partially responsible for vascular side effects that occur in the management of diabetes. These side effects may be overcome by the development of a more physiologic route of administration than by subcutaneous injection.

Insulin in the Drinking Water of Rats

AbstractThis work was supported by a grant from the Defiance Area Diabetes Club. We acknowledge with thanks the gift of human insulin from Novo-Nordisk, Denmark. We thank G. Colin Budd and Ben Pansky for their intellectual contributions and encouragement. We thank Robert J. Trumbly for his hospitality in his laboratory and for his financial support of FEW. We thank R. Birkhahn for the use of metabolic cages in this project and the personnel of the Laboratory Animal Research Facility for their expert care of the rats.A preliminary account of this work was presented at a Symposium of the Controlled Release Society in Dublin, 20–22 September 1995 [5].Normal and streptozotocin-diabetic female Sprague–Dawley rats were given solutions of insulin to drink instead of water. Tail vein blood glucose concentrations were monitored by Glucometer. Urine volume, food and water intake, and body weight were recorded daily. Diabetic rats exhibited decreased daily urinary output when more than 70 IU/mL of insulin was in the...

Properties of CRF from Normal and Brattleboro Rat Median Eminence

Most of the corticotrophin-releasing factor (CRF) activity of normal rat median eminence (ME) extract binds to a neurophysin affinity column. The bound material contains the large and small factors, which we have previously demonstrated to be required together for full activity. Most of the CRF activity of Brattleboro rat ME extract, which contains as much CRF activity as the ME extract of a normal rat, does not bind to a neurophysin affinity column. The CRF activity of Brattleboro rat ME extract resides entirely in a large molecule as determined by Sephadex G-25 chromatography. The different properties of Brattleboro and normal rat CRF suggest that the CRF activity in the Brattleboro rat may result from a substance which is different from that in a normal rat.