Paul G. Wood

Stress-induced alterations in interferon production and class II histocompatibility antigen expression ☆

Mild electric foot-shock has been shown to be a stressor that can alter immune responses. Male Lewis rats were exposed to one session of 16 5.0-s 1.6-mA foot-shocks. Production of interferon-gamma by splenocytes in response to concanavalin-A was decreased in spleens from the shocked rats compared to control spleens. Spleen cells from rats treated with nadolol, a peripherally acting beta-adrenergic receptor antagonist, and then shocked, showed dose-dependent attenuation of the suppression of interferon-gamma production. This suggests that catecholamines mediate shock-induced suppression of interferon-gamma production. The percentage of splenic mononuclear cells expressing class II histocompatibility (Ia) antigens on their surfaces from spleens of shocked rats was determined by flow cytometry. Significantly decreased class II positive mononuclear cells were present in the spleens of shocked rats in comparison to the spleens of control rats. This may reflect an alteration of cell trafficking or decreased production of class II antigens.

2-deoxy-D-glucose modulation of T-lymphocyte reactivity: differential effects on lymphoid compartments.

This study was designed to evaluate the effect of glucoprivation, as induced by 2-deoxy-D-glucose (2-DG) administration, on lymphocyte mitogen reactivity in Sprague-Dawley rats. The results showed that a single injection of 2-DG decreased reactivity in both whole-blood and spleen lymphocytes, as determined by mitogenic stimulation to concanavalin A (Con A) and phytohemagglutinin (PHA). However, the suppressed reactivity for the spleen lymphocytes attenuated with repeated injections, but the whole-blood lymphocytes did not show attenuation. Mitogen assessments of lymphocytes obtained from the thymus indicated that a single injection did not induce suppressed reactivity, but repeated injections induced a pronounced suppression of responsiveness. Furthermore, mitogen assessments of mesentery lymph nodes did not show any effect of 2-DG injections. These results corroborate other findings using electric shock as the stressor, namely that different compartments of the immune system are differentially affected by a stressor.

The area postrema lesions alter the inhibitory effects of peripherally infused pancreatic polypeptide on pancreatic secretion

Circulating PP binds to specific receptors in the DVC through the AP, but the mechanism through which these brain receptors affect pancreatic secretion is not clear. We hypothesize that the removal of the AP (APX) will alter the effects of PP on pancreatic secretion. APX or sham procedures were performed in anesthetized male Wistar rats. After a 1-month recovery, one group of rats were infused with either PP (30 or 100 pmol/kg per h) or vehicle under basal or 2-DG-stimulated (75 mg/kg, i.v. bolus) conditions for studying pancreatic exocrine secretion. A second parallel group was sacrificed for examination of PP receptor binding in the brain stem. A third group received an intraperitoneal injection of PP at the dose of 4.15x10(4) pmol/kg (200 microg/kg) and c-fos expression in the brain stem was examined. APX eliminated PP binding sites in the DVC as assessed by autoradiography. PP infusion caused a dose-dependent decrease in basal protein secretion. APX partially reversed PP inhibition of basal protein secretion when infused at 30 pmol/kg per h, and at 100 pmol/kg per h stimulated pancreatic fluid secretion and reversed the inhibition of protein secretion. During 2-DG stimulation the effects of PP and 2-DG on pancreatic fluid and protein secretion were parallel. PP dose-dependently inhibited 2-DG-stimulated secretion in sham rats. APX reduced the pancreatic fluid (54%) and protein (46%) secretory response to 2-DG. However, PP at 30 pmol/kg per h remained a potent inhibitor of 2-DG-stimulated pancreatic secretion in APX rats. This effect was blunted with PP at 100 pmol/kg per h in APX rats, possibly related to the stimulatory effect of high-dose PP in APX rats without 2-DG. Furthermore, i.p. PP induced significantly greater c-fos activation of NTS neurons in APX rats than sham rats, despite the apparent absence of PP binding sites in the DVC. We conclude that in awake rats, PP inhibits basal secretion, in part, through the AP. Furthermore, and unlike PYY, PP inhibits 2-DG-stimulated pancreatic secretion, and it does so through an AP-independent mechanism. The possibility that the mechanism may involve the DVC cannot be excluded since i.p. injection of PP activates c-fos expression in DVC neurons. Thus, PP and PYY may regulate different components of the pancreatic secretory control system through unique pathways.

Chronic Alcohol-Induced Alterations in the Pancreatic Secretory Control Mechanisms

Chronic alcohol ingestion appears to increase susceptibility of the pancreas to pancreatitis through multiple mechanisms. The aim of the current study was to determine the effect of chronic low- and high-dose alcohol consumption on the neurohormonal control of the exocrine pancreas in rats. Male Wistar rats were fed Lieber DeCarli liquid control-, low-, and high-dose alcohol diets for 3 months. Pancreatic exocrine secretion was measured under basal and 2-deoxy-d-glucose (2-DG)-, CCK-, bethanechol-, or meal-stimulated conditions while on chronic alcohol diets and after 2-DG or CCK stimulation during alcohol withdrawal in awake rats. Chronic alcohol ingestion was associated with a dose-related inhibition of basal pancreatic protein secretion, which was reversed upon alcohol withdrawal. Low-dose alcohol feeding had no effect on bethanechol-stimulated pancreatic secretion but altered 2-DG-stimulated pancreatic secretion. In chronic high-dose alcohol rats, meal- and bethanechol-stimulated protein secretion was significantly potentiated during early and late phases. The response to CCK appeared to be disinhibited, whereas the response to 2-DG was uniformly blunted. Upon withdrawal of low-dose alcohol, the response to 2-DG was potentiated, whereas with the withdrawal of high-dose alcohol, the response to CCK was potentiated. Adaptation to chronic alcohol consumption differs depending on the alcohol dose. The most significant effects were seen after high-dose alcohol withdrawal, with apparent loss of central inhibitory regulation combined with exaggerated response at the acinar cell level. This combination of factors could increase susceptibility to acute alcoholic pancreatitis through a hyperstimulation mechanism.

Cellular distribution of secretin receptor expression in rat pancreas

Secretin is an important regulator of pancreatic function, but the molecular basis of its actions is not well understood. We have, therefore, used in situ autoradiography, photoaffinity labeling, and RNase protection assays with healthy rat pancreas, dispersed acinar cells, and pancreas depleted of acinar cells to explore the cellular distribution and molecular identity of high-affinity secretin receptors in this complex organ. The autoradiographic examination of125I-labeled [Tyr10]rat secretin-27 binding to normal pancreas demonstrated saturable and specific high-affinity binding sites on both acinar and duct cells, with a uniform lobular distribution, but with no binding above background over islets or vascular structures. Photoaffinity labeling demonstrated that the ductular binding site in acinar cell-depleted copper-deficient rat pancreas represented the same glycoprotein with a molecular weight of 50,000-62,000 that was present on acinar cells. RNase protection assays confirmed the molecular identity of the secretin receptors expressed on these distinct cells. The apparent absence or extreme low density of similar secretin receptors on islets and pancreatic vascular structures suggests that the pharmacological effects of secretin on those cells may either be indirect or mediated by another secretin family receptor that recognizes this hormone with lower affinity.Secretin is an important regulator of pancreatic function, but the molecular basis of its actions is not well understood. We have, therefore, used in situ autoradiography, photoaffinity labeling, and RNase protection assays with healthy rat pancreas, dispersed acinar cells, and pancreas depleted of acinar cells to explore the cellular distribution and molecular identity of high-affinity secretin receptors in this complex organ. The autoradiographic examination of 125I-labeled [Tyr10]rat secretin-27 binding to normal pancreas demonstrated saturable and specific high-affinity binding sites on both acinar and duct cells, with a uniform lobular distribution, but with no binding above background over islets or vascular structures. Photoaffinity labeling demonstrated that the ductular binding site in acinar cell-depleted copper-deficient rat pancreas represented the same glycoprotein with a molecular weight of 50,000-62,000 that was present on acinar cells. RNase protection assays confirmed the molecular identity of the secretin receptors expressed on these distinct cells. The apparent absence or extreme low density of similar secretin receptors on islets and pancreatic vascular structures suggests that the pharmacological effects of secretin on those cells may either be indirect or mediated by another secretin family receptor that recognizes this hormone with lower affinity.

Corticosterone-independent alteration of lymphocyte mitogenic function by amphetamine.

Amphetamine, a neural stimulatory agent with acute effects mimicking those of stress, is shown here to elevate plasma corticosterone levels and suppress spleen and peripheral blood lymphocyte (PBL) mitogenic responses to concanavalin A (Con A) and phytohemagglutinin (PHA) when administered to rats. Pretreatment of the rats with propranolol, a nonselective beta-adrenergic receptor antagonist, totally prevented the amphetamine-induced suppression of lymphocyte mitogenic reactivity to Con A and PHA in the spleen and to PHA in the peripheral blood; however, the PBL mitogenic response to Con A was only partially restored. Although the amphetamine-induced alterations in immune function were prevented by propranolol pretreatment, the elevated plasma corticosterone response was not. This suggests that corticosterone is not modulating the mitogenic activity of splenic lymphocytes or PHA-reactive PBLs. On the other hand, Con A-reactive PBLs may be affected by corticosterone and/or other mechanisms, which may include the catecholamines.

β2-Adrenergic receptor density and cardiovascular response to mental stress

In this study we evaluated effects of an acute experimental stressor on beta 2-adrenoceptor density and examined the relationships of baseline receptor density to cardiovascular reactions induced by stress. In addition, we investigated whether any observed alterations in receptor density were associated with concomitant redistribution of circulating lymphocyte populations. Receptor density and lymphocyte subsets were determined before and immediately following performance of a frustrating laboratory task in 22 male volunteers. Blood pressure, heart rate (HR), and plasma catecholamine concentrations were also assessed at baseline and during task performance. Parallel measurements were obtained among 11 unstressed control subjects. Receptor density increased significantly between baseline and posttask measurements, but equally so in experimental and control subjects. Numbers of T suppressor/cytotoxic and natural killer cells increased selectively among subjects assigned to the experimental (stress) condition. However, there was no association between lymphocyte subset distribution and receptor density. Interindividual variability in pretask receptor density correlated significantly with heart rate and systolic blood pressure (SBP) reactivity during the initial 3 min of mental stress, but not over the entire task period. In addition, baseline receptor density correlated with SBP (but not HR) reactivity after covariance adjustment for the concomitant change in plasma catecholamine concentrations.

PYY Potently Inhibits Pancreatic Exocrine Secretion Mediated Through CCK-Secretin-Stimulated Pathways but Not 2-DG-Stimulated Pathways in Awake Rats

Peptide YY (PYY) is an important modulator of stimulated pancreatic exocrine secretion. PYY acts proximal to the acinar cell but the exact site and mechanism of action are unknown. The aim of the present study is to determine the pathway through which PYY exerts its effect on the exocrine pancreas in awake rats under physiological condition. When pancreatic secretion was stimulated by graded doses of cholecystokinin (CCK) (14, 28, 58 pmol/kg/hr) with secretin (1.25, 2.5, 5.0 pmol/kg/hr) or CCK alone at 28 pmol/kg/hr, PYY 1-36 dose-dependently inhibited pancreatic secretory responses. Moreover, PYY1-36 at 50 pmol/kg/hr almost completely blocked the stimulation by CCK (P < 0.01). Although background infusion of PYY1-36 or PYY3-36 at 12.5 pmol/kg/hr inhibited basal pancreatic fluid and protein secretion, but both of them only partly inhibited the subsequent 2-DG stimulated pancreatic fluid and protein secretion. Furthermore, PYY1-36 at 50 pmol/kg/hr failed to inhibit 2-DG-stimulated pancreatic secretion. These results confirm that PY1-36 inhibits CCK-stimulated pancreatic secretion under all experimental conditions. However, in the awake, surgically recovered rat, PYY1-36 at both low and high doses failed to fully inhibit 2-DG-stimulated pancreatic secretion. Therefore, the site of PYYs inhibitory action on pancreatic secretion appears to be primarily on the CCK-stimulated pathway at a site proximal to the convergence of the CCK and 2-DG pathways.

PYY Regulates Pancreatic Exocrine Secretion Through Multiple Receptors in the Awake Rat

Peptide YY (PYY) is one of several regulatory peptides reported to modulate pancreatic secretion. PYY circulates in two forms, PYY1–36 and PYY3–36, and binds to multiple receptor subtypes. We sought to determine if PYY1–36 or PYY3–36 regulates neurally mediated pancreatic secretion through the Y1, Y2, and/or Y5 receptor subtypes. Experiments were conducted in awake, surgically recovered rats. In order to determine the effects of the PYYs on basal pancreatic secretion, either PYY1–36, [Pro34] PYY1–36 (a Y1/Y5 agonist), or PYY3–36 (a Y2/Y5 agonist) were infused for 40 min at doses of 0, 12.5, 25, or 50 pmol/kg/hr while measuring pancreatic juice volume and protein. PYY1–36 increased pancreatic protein secretion at 25 and 50 pmol/kg/hr (P < 0.05) in a dose-dependent manner (P < 0.001, R2= 0.990). The Y2/Y5 receptor agonist PYY3–36 significantly inhibited pancreatic juice volume and protein at 12.5 and 25 pmol/kg/hr, but stimulated protein secretion at higher doses (P < 0.001, R2= 0.995). The Y1/Y5 agonist, [Pro34] PYY1–36, had no significant effect on basal pancreatic exocrine secretion. Therefore, PYY1–36, PYY3–36 and [Pro34] PYY1–36 produced different, dose-dependent changes on basal pancreatic exocrine secretion. Inhibition of pancreatic secretion by circulating PYY1–36 and PYY3–36 are primarily mediated by the Y2 receptor. Since [Pro34] PYY1–36 did not change pancreatic secretion, it can be concluded that circulating PYY1–36 or PYY3–36 does not modulate pancreatic secretion through the Y1 or Y5 receptors. Since the stimulatory effects of PYY1–36 on pancreatic secretion could not be explained by the actions of PYY3–36 or [Pro34] PYY1–36 on Y1 or Y2 receptors, and since PYY1–36 fails to bind to Y3 or Y4 receptors, we also conclude that PYY1–36 may stimulate pancreatic secretion in a dose-dependent mechanism through a PYY receptor subtype different from Y1, Y2, Y3, Y4 or Y5.

Area Postrema Lesion Alters the Effects of Peptide YY on 2-DG-Stimulated Pancreatic Exocrine Secretion

Previously we demonstrated that circulating peptide YY (PYY), which inhibits pancreatic exocrine secretion, binds to specific receptors in the area postrema (AP); therefore we have tested the hypothesis that the removal of the AP (APX) will alter the effects of PYY on pancreatic secretion in awake rats. One-month after AP lesion or sham lesion, rats were implanted with pancreatic, biliary, duodenal, and intravenous catheters. After recovery from the surgery, unanesthetized rats were infused with vehicle or PYY (30 pmol/kg/hr or 100 pmol/kg/hr) under basal or 2-deoxy-d-glucose (2-DG) stimulated (75 mg/kg, intravenous bolus) conditions. PYY at 30 pmol/kg/hr inhibited basal pancreatic fluid secretion in sham-operated rats, but not APX rats. PYY at 100 pmol/kg/hr stimulated basal pancreatic protein secretion in sham-operated rats, and this effect was also lost in APX rats. PYY at 30 and 100pmol/kg/hr inhibited peak 2-DG stimulated protein secretion to a greater extent in APX rats as compared to sham-operated rats (P < 0.05). Since PYY inhibition of basal pancreatic secretion is AP dependent and inhibition of 2-DG stimulated pancreatic secretion is AP independent, we conclude that the 2-DG pathway of pancreatic secretion differs from the pathway responsible for basal secretion, and that APX potentiates the inhibitory effect of PYY on the 2-DG pathway.