Catechol conjugation with hemolymph proteins and their incorporation into the cuticle of the American cockroach, Periplaneta americana.

Newly ecdysed American cockroaches, Periplaneta americana (sixth to last instar) were injected with radioactive dopamine (DA) and hemolymph was collected at 10-60 min post-ecdysis. Size-exclusion chromatography established the presence of at least three proteins that serve as catecholamine carriers. Reinjection of the smaller radiolabeled phenol-bound proteins into newly ecdysed animals results in in vivo aggregation, with the radiolabel bound to large MW proteins (30- > 200 kDa). In addition, the reinjection of radiolabeled protein of any size resulted in the incorporation of the label into the newly sclerotized cuticle. Hemolymph proteins were synthesized in vivo using [14C]leucine and subsequently double labeled in vivo with [3H]dopamine. After sclerotization (7 h post-ecdysis) the cuticle was extirpated, hydrolyzed and counted. An identical ratio of 14C to 3H was found in cuticle extracts as in the double-labeled hemolymph proteins, suggesting that the phenol-bound protein was incorporated in the cuticle unchanged. It appears that the catechol bound to the proteins exists as a beta-glucoside.

Effects of trichothecene mycotoxins on the ex vivo vascular smooth muscle responses to beta-adrenergic agonists.

The ex vivo biophysical response of isolated rat aorta to T-2 toxin, diacetoxyscirpenol (DAS) and Roridin-A were determined. Roridin-A caused a dose-dependent relaxation of precontracted (30 mM KCl) aorta, while T-2 toxin and DAS were without effect. On the other hand, the relaxation response of aorta to isoproterenol and sodium fluoride is impaired in the presence of T-2 toxin or DAS. Roridin-A did not alter the relaxation response to isoproterenol or sodium fluoride. These experiments suggest that macrocyclic trichothecenes (e.g., Roridin-A) are capable of altering vascular smooth muscle tone directly, while simple trichothecenes (e.g., T-2 toxin, DAS) interfere with beta-adrenergic responses.

Urea kinetics in humans at two levels of exercise intensity.

A primed constant infusion of [15N2]urea was used to quantify the response of urea production to exercise at 40 and 70% maximal oxygen consumption on a treadmill. Total urea production, urea production from recycled N, urea production from nonrecycled N, and urea N recycled back into body protein were calculated. Most components of urea kinetics were unaffected by exercise at either intensity. The rate of urea reincorporated into protein was significantly increased during exercise and recovery at both levels of exercise. We conclude that exercise does not stimulate urea production but that there may be an accelerated reincorporation of urea N back into body protein.

The effect of aflatoxin B1 exposure on serotonin metabolism: response to a tryptophan load.

Semi-chronic exposure of ICR male Mice to Aflatoxin B1 (AFB1) in non-toxic doses decreased brain serotonin (5-hydroxytryptamine, 5-HT), 5-hydroxyindole-3-acetic acid (5-HIAA) and catecholamines without altering tryptophan (TRP) or tyrosine (TYR) levels. A TRP load (300 mg/kg, i.p. x 2 hours) slightly increased brain TRP levels while causing a slight decrease in 5-HT and 5-HIAA in control animals. A TRP load in AFB1 treated mice increased brain 5-HT and 5-HIAA. The TRP load caused a further reduction in brain catecholamines without altering TYR levels. Exposure to AFB significantly increased lung TRP levels without altering 5-HT or 5HIAA levels. TRP loading increased lung TRP concentrations in control mice. However, in AFB1 treated mice the increase was not significantly elevated above the level caused by AFB1 treatment alone. Lung 5-HT or 5-HIAA levels in control or AFB1 treated mice are not significantly altered by TRP loading. These experiments demonstrate that AFB1 alters brain and lung TRP metabolism.

Identifying body fluid distribution by measuring electrical impedance.

The effects of critical illness on extracellular water (ECW) and total body water (TBW) were measured using (1) a multiple dilutional technique, and (2) whole body and regional bioelectrical impedance analysis (BIA) in a group of stable patients. Total body water and body resistance (R) were similar in patients when compared with normal healthy subjects (TBW: 45.1 +/- 4.5 vs. 46.2 +/- 3.4 L, p = 0.85; R: 518 +/- 42 vs. 500 +/- 22 omega, p = 0.70), and a significant relationship was present between these measurements (r = -0.87, p < 0.001). However, patients demonstrated an increase in ECW compared with controls (ECW: 18.6 +/- 1.3 vs. 14.7 +/- 1.1 L, p < 0.05). Expanded ECW values were associated with diminished electrical reactance (Xc) values (38 +/- 6 vs. 70 +/- 4 omega, p < 0.001) and these values were correlated (r = -0.67, p < 0.005). The ratio of Xc to R determined across the body and each of the segments was significantly lower in patients compared with controls (at least p < 0.005) and this ratio measured across a leg was the most sensitive predictor of health (Xc/R > or = 0.137) and disease (Xc/R < or = 0.101). Bioelectrical impedance analysis is a noninvasive and simple bedside technique that can be used to predict TBW and identify altered fluid distribution following critical illness.

Growth hormone attenuates the abnormal distribution of body water in critically ill surgical patients.

BACKGROUND: Catabolic illness is associated with fluid retention and extracellular space expansion. To determine the effect of human growth hormone (GH) on body water compartments, critically ill surgical patients were studied for a 2-week period during which they either continued to receive standard intensive care unit support, or in addition, received GH, 10 mg/day. METHODS: Body water compartments were measured at the beginning and end of the period by the indicator dilution technique with sodium bromide and heavy water used as the indicators of extracellular (ECW) and total body water (TBW), respectively; intracellular water (ICW) was calculated by subtraction. RESULTS: Neither group lost significant amounts of weight or TBW. A marked ECW expansion and disturbance of the ECW/TBW ratio occurred in the patients receiving standard care, which was associated with a dramatic reduction in ICW, a critical component of the body cell mass (BCM). In contrast, GH-treated patients maintained ECW and ICW, indicating a preservation of BCM, and their ECW/TBW ratio normalized. CONCLUSIONS: GH administration prevents ECW retention and stabilizes or normalizes fluid distribution during critical illness. Taken together with its known anabolic effects under these conditions, the maintenance of ICW demonstrates that GH can be used to preserve BCM in complex surgical patients.

Insulin-like growth factor-I response is comparable following intravenous and subcutaneous administration of growth hormone.

Subcutaneous (sq) administration of recombinant human growth hormone (r-hgh) has an anabolic effect and increases systemic insulin-like growth factor (IGF-I) in surgical patients. IGF-I is a mediator of growth hormone (gh) anabolic effects. To determine the effect of intravenous (iv) administration of r-hgh on systemic IGF-I, 11 patients were given 14 1-week courses of daily 8-hr infusions of r-hgh (10 mg in 500 ml D5W). Serum gh and IGF-I levels were measured. To compare routes of administration, iv r-hgh patients were matched to comparable sq r-hgh patients and IGF-I responses were examined. Illness severity effect on IGF-I response to r-hgh was assessed by dividing 16 burn patients who received either iv or sq r-hgh into two groups on the basis of severity scores. Analysis of the data showed that IGF-I levels increased significantly after iv r-hgh, IGF-I response to iv r-hgh (1.14 +/- 0.18 U/ml to 4.12 +/- 0.65 U/ml) was not different from IGF-I response to sq r-hgh (1.04 +/- 0.36 U/ml to 4.96 +/- 1.09 U/ml). Increasing illness severity attenuated the IGF-I response in the more severely injured group (0.91 +/- 17 U/ml to 2.40 +/- 0.38 U/ml) relative to the less severely injured group (1.37 +/- 0.22 U/ml to 5.53 +/- 0.78 U/ml) despite a significant increase in IGF-I after gh in both groups. In summary, IGF-I increased significantly after iv r-hgh and the increases were similar to those seen after sq r-hgh in comparable patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Alterations in body fluid content can be detected by bioelectrical impedance analysis.

The electrical resistance across the whole body and its segments to the conduction of a weak alternating current was determined in human subjects under three different conditions: (1) during bed rest, (2) during infusion of 1 liter of saline, and (3) during donation of 1 unit of blood. During bed rest, extracellular and total body water were measured by dilution of bromide and heavy water, respectively. Electrical resistance obtained from electrodes placed on proximal portions of extremities ("proximal resistance") accounted for less than 50% of that determined by electrodes positioned on routinely used portions of a hand and foot ("whole body resistance"). Following saline infusion, resistance determined from the whole body and all its segments fell (P less than 0.001); the magnitude of the drop in both proximal and whole body resistance was inversely related to the volume of total body water (TBW) (r = -0.82, P less than 0.002, and r = -0.73, P less than 0.01, respectively). In contrast, blood donation was associated with significantly increased resistance at both measurement sites. TBW predicted from anthropometrics was inversely related to both proximal (r = -0.90, P less than 0.001) and whole body resistance (r = -0.75, P less than 0.001). Bioelectrical impedance analysis is a simple technique which may be useful in monitoring minimal alterations in TBW. Furthermore, altered fluid status may be predicted more accurately by changes in proximal resistance compared to changes in traditionally used whole body resistance.

Acute and chronic effects of the trichothecene mycotoxin T-2 on rat brain regional concentrations of serotonin, tryptophan, and tyrosine.

Female rats were given 1 acute dose or chronic doses (once every 48 hr for 28 days) of T-2 toxin (10 micrograms/kg ip) or vehicle. At necropsy, each brain was subdivided into cerebellum, cerebral cortex (including telencephalon and diencephalon), and brainstem (including mesencephalon, metencephalon, and myelencephalon). Acute systemic T-2 toxin administration increased cerebellar and brainstem tryptophan while serotonin, a tryptophan metabolite, was decreased correspondingly in these same brain regions. Chronic T-2 administration increased cerebellar tyrosine and serotonin concentrations, while cortical tryptophan concentrations were also increased. These results indicate that both acute and chronic administration of T-2 toxin cause differential changes in regional distribution levels of tyrosine, tryptophan, and serotonin.

Differential changes in rat brain tryptophan, serotonin and tyrosine levels following acute aflatoxin B1 treatment.

Female Sprague-Dawley rats were treated acutely (12-h) with aflatoxin B1 (100 micrograms/kg i.p.) or vehicle (10% acetone in 0.9% NaCl) and regional brain levels of tryptophan, serotonin and tyrosine were assayed. Brainstem but not cerebellar or cortical tyrosine levels were decreased in aflatoxin B1-treated rats. Brain tryptophan was increased in all 3 brain regions by acute aflatoxin B1 treatment, while serotonin levels were unaltered in the cerebellum and cortex and decreased in the brainstem. These experiments indicate that acute aflatoxin B1 treatment differentially alters brain amino acids and serotonin and that changes in brain tryptophan, the serotonin precursor, do not parallel changes in brain serotonin.

Disturbances in tryptophan metabolism in rats following chronic dietary aflatoxin treatment.

Male Holtzman rats were fed 7.8 ppm mixed aflatoxins (AFT) for a 21-week period. At sacrifice there was extensive bile ductule cell proliferation and numerous precancerous changes evident in AFT-treated animals. The in vitro activity of hepatic holo-enzyme tryptophan 2, 3-dioxygenase was reduced in AFT-treated animals which correlated with a reduced urinary excretion of both kynurenine and kynurenic acid suggesting that this reduced tryptophan 2, 3-dioxygenase observed in vitro may be relevant to the in vivo response. The levels of serotonin and 5-hydroxyindole-3-acetic acid were found to be reduced in the pineal, pituitary, various brain regions and in duodenum of AFT-treated rats. It is proposed that the decreased levels of serotonin and 5-hydroxyindole-3-acetic acid in all tissues may be due to either a decreased free tryptophan pool or alterations in 5-hydroxyindole biosynthesis.

Disturbances of tryptophan metabolism in mice acutely deprived of tryptophan.

The effect of a dietary tryptophan deficiency on tissue serotonin (5-hydroxytryptamine) synthesis rates, systemic tryptophan metabolism and its response to steroid or cycloheximide treatment was investigated. Brain serotonin synthesis was depressed in tryptophan-deprived (TD) mice while duodenal serotonin synthesis was enhanced following a tryptophan load. Liver total protein content was initially depressed in TD mice but increased following a tryptophan load. Blood tryptophan and albumin were depressed in TD mice while the percentage of albumin-bound tryptophan significantly increased in TD mice. Serum-free fatty acids were not significantly altered. Furthermore, enzyme kinetics studies indicated that in TD mice, tryptophan-5-hydroxylase has a reduced Vmax, while the Km for both tryptophan and the pteridine cofactor was significantly lowered. The tryptophan hydroxylase response to tryptophan or hydrocortisone injection was accentuated in TD mice while the tryptophan-2,3-dioxygenase response to tryptophan or hydrocortisone injection was blunted in TD mice. Finally, injection of tryptophan and cycloheximide blocked the tryptophan-2,3-dioxygenase response while the tryptophan hydroxylase response was unaltered in both control and TD diet mice.

The effect of a high-fat diet on brainstem and duodenal serotonin (5-HT) metabolism in Sprague-Dawley and Osborne-Mendel rats.

Sprague-Dawley (S-D) and Osborne-Mendel (O-M) rats were fed either a low-fat diet (5 percent corn oil) or high-fat diet (20 percent corn oil) for a six-week-period. Brainstem and duodenal levels of tryptophan, serotonin (5-hydroxytryptamine, 5-HT) and 5-hydroxyindole-3-acetic acid (5-HIAA) were not altered by dietary treatment in the O-M rats. On the other hand, the high-fat diet significantly decreased brainstem 5-HT levels in S-D rats. Brainstem and duodenal 5-HT levels were decreased in O-M rats as compared to S-D rats and this phenomenon is not altered by dietary treatment. It is suggested that the O-M rat may have a alteration in the 5-HT metabolic system and that such a defect may contribute to the development of obesity.

Reversal of acute streptozotocin-induced glycemic changes by chemical ventro-medial hypothalamic lesions.

This report describes the effect of chemical gold thioglucose (GTG) lesions of the ventromedial hypothalamic (VMH) nuclei in male Sprague-Dawley rats previously made diabetic by injection of streptozotocin (50 mg/kg, i.v.). Injection of streptozotocin resulted in hyperglycemia (blood glucose greater than 200 mg%) within one week; treatment of these rats with intracerebroventricular (ICV) injections of GTG (0.025 M) + Insulin (1 microU) reversed these glycemic changes within six weeks. On the basis of these initial experiments, it has been shown that GTG lesions of the VMH may reverse glycemic changes in acute streptozotocin-induced diabetes.

Alterations of tryptophan metabolism in a rat strain (Osborne-Mendel) predisposed to obesity.

1. Osborne-Mendel (O-M) rats displayed differences in brain and systemic tryptophan metabolism. O-M rats had decreased brainstem tryptophan-5-hydroxylase activity and decreased serotonin (5-HT) levels as compared to Sprague-Dawley rats. However, brain tryptophan levels were actually increased in O-M rats. Norepinephrine, dopamine and 5-hydroxyindole-3-acetic acid levels were not different between strains. 2. Pineal serotonin levels were increased in O-M rats. 3. Liver tryptophan 2,3-dioxygenase activity was increased in O-M rats while tyrosine aminotransferase activity was not different between strains. 4. Total blood cholesterol was decreased in O-M rats while triglycerides, free fatty acids and albumin was not different between strains. Total serum tryptophan was not different between strains while O-M rats had an increased level of free (unbound) tryptophan.

Dietary aflatoxin and copper acetate effects on various blood parameters in rats.

The effects of copper acetate (CuAc) and aflatoxin (AFT) dietary supplements on serum vitamins A and E, albumin, cholesterol, and the formed elements of blood in male rats were investigated. The CuAc and AFT dietary supplements were 0.5% and 7.8 ppm respectively. Vitamins A and E, red blood cell count (RBC), mean cell volume (MCV), mean cell hemoglobin content (MCHC) and hematocrit (Hct) were not significantly altered by either supplement while albumin and cholesterol was significantly elevated in groups receiving either CuAc or AFT. Serum hemoglobin (HBG) was depressed in animals receiving the AFT + CuAc supplement while the WBC count was depressed in both groups receiving CuAc while AFT had no effect.

Alterations of brain and intestine serotonin levels in hamsters pretreated with dietary aflatoxin.

Male Syrian hamsters were fed 30.3 ppm mixed aflatoxins (AFT) for a 45-day period followed by a 123-day recovery period. Tests made at the end of the recovery period indicated a reduction in whole brain and duodenal serotonin and a slight but not statistically significant elevation in brain 5-hydroxyindoleacetic acid. There was a high incidence of liver cell hyperplasia in the aflatoxin fed animals. Following toxin withdrawal, the experimental animals showed significant gains in body weight but they never attained a body weight equal to the controls. Aflatoxin apparently interferes with serotonin production and the return to normal body weight even after withdrawal of the animal from the contaminated diet.