Significance of tissue myo-inositol concentrations in metabolic regulation in nerve.

Approximately 25 percent of resting energy utilization in isolated nerve endoneurium is inhibited by medium containing defatted albumin and selectively restored by arachidonic acid but is unaffected by indomethacin or nordihydroguaiaretic acid. The same component of energy utilization is inhibited by small decreases in endoneurial myo-inositol, which decrease incorporation of carbon-14-labeled arachidonic acid into phosphatidylinositol. The fraction of the resting oxygen uptake inhibited by ouabain is decreased 40 to 50 percent by a reduced tissue myo-inositol concentration or by defatted albumin. Metabolic regulation by rapid, basal phosphatidylinositol turnover is dependent on the maintenance of normal tissue myoinositol concentrations.

Basal phosphatidylinositol turnover controls aortic Na+/K+ ATPase activity.

To determine whether basal phosphoinositide turnover plays a role in metabolic regulation in resting rabbit aortic intima-media incubated under steady state conditions, we used deprivation of extracellular myo-inositol as a potential means of inhibiting basal phosphatidylinositol (PI) synthesis at restricted sites and of depleting small phosphoinositide pools with a rapid basal turnover. Medium myo-inositol in a normal plasma level was required to prevent inhibition of a specific component of basal de novo PI synthesis that is necessary to demonstrate a discrete rapidly turning-over [1,3-14C]glycerol-labeled PI pool. Medium myo-inositol was also required to label the discrete PI pool with [1-14C]arachidonic acid (AA). The rapid basal turnover of this PI pool, when labeled with glycerol or AA, was not attributable to its utilization for polyphosphoinositide formation, and it seems to reflect basal PI hydrolysis. Depleting endogenous free AA with medium defatted albumin selectively inhibits the component of basal de novo PI synthesis that replenishes the rapidly turning-over PI pool. A component of normal resting energy utilization in aortic intima-media also specifically requires medium myo-inositol in a normal plasma level and a free AA pool; its magnitude is unaltered by indomethacin, nordihydroguaiaretic acid, or Ca2+-free medium. This energy utilization results primarily from Na+/K+ ATPase activity (ouabain-inhibitable O2 consumption), and in Ca2+-free medium deprivation of medium myo-inositol or of free AA inhibits resting Na+/K+ ATPase activity to a similar degree (60%, 52%). In aortic intima-media basal PI turnover controls a major fraction of resting Na+/K+ ATPase activity.

Effects of polyol-pathway inhibition and dietary myo-inositol on glomerular hemodynamic function in experimental diabetes mellitus in rats.

Early functional disturbances in nerve, retina, and lens in diabetes mellitus appear to result from a common mechanism involving increased polyol-pathway activity with an associated effect on tissue myo-inositol metabolism. We tested the role of increased polyol-pathway activity in the early glomerular hemodynamic abnormalities in experimental diabetes in rats with dietary myo-inositol supplementation or the administration of sorbinil, an aldose reductase inhibitor. Each maneuver prevented the glomerular hyperfiltration of early streptozocin-induced diabetes and reversed the hyperfiltration of established diabetes of 10 days' duration. We also found that the abnormal response to captopril in diabetic rats was improved by dietary myo-inositol supplementation or sorbinil administration. Although nonhypotensive doses of captopril lowered glomerular filtration rate (GFR) in diabetic rats on a 0.01% myo-inositol diet, GFR increased substantially after captopril infusion in diabetic rats treated with sorbinil or myo-inositol supplementation. These data suggest that normalization of tissue myo-inositol metabolism restores normal responsiveness to angiotensin II; this may contribute to the reduction in GFR with the two experimental maneuvers. We also tested the interaction between polyol-pathway activation and high dietary protein intake. Aldose reductase inhibition and dietary myo-inositol supplementation had no effect on the component of increased GFR due to 50% dietary protein intake but specifically inhibited the hyperfiltration attributable to diabetes. These results suggest that hyperglycemia acts through increased polyol-pathway activity and its effects on tissue myo-inositol metabolism to play a fundamental role in the pathogenesis of the glomerular hyperfiltration characteristic of early diabetes.

Protein expression in relation to the cell cycle of exponentially growing human prostatic epithelial cells.

This report concerns the study of the relationship between protein expression and the cell cycle in exponentially proliferating benign and malignant human prostate epithelial cells in short-term cultures. Multiparameter flow cytometric measurements were performed to correlate the expression of prostate-specific acid phosphatase, epithelial membrane antigen and epitectin with cell cycle progression. The expression of the three proteins was heterogeneous in G1 cells. The early post-mitotic cells exhibited the lowest levels when compared with late G1 cells, wherein the expression was many times greater. There was no further increase as the cells progressed through S and G2 + M. These findings, corroborating prior observations in other systems, suggest the possibility that the levels of the proteins studied increase during the G1 phase of the cell cycle and drop during or immediately after cytokinesis. As an alternate explanation, the heterogeneity of protein expression characteristic of G1 cells may be due, at least in part, to an asymmetric apportionment of cell constituents at mitosis.

Forebrain expression of c-fos due to active maternal behaviour in lactating rats.

To reveal brain sites simultaneously active during the expression of maternal behaviour in lactating rats, we used immunocytochemical visualization of the nuclear protein product Fos of the immediate-early gene c-fos as a marker of neuronal activity. After a 48 h separation from their litter, day 7 postpartum dams received a 1 h period of physical interaction with pups either capable or incapable of suckling, inaccessible pups in a wire-mesh box, an empty box, or no stimulation. Physical interaction with pups elicited high levels of pronurturant maternal behaviour (retrieval, licking, mouthing), and suckling elicited nursing behaviour as well. Exposure to the box, with or without pups, elicited high levels of investigatory sniffing, self-grooming, and general activity. Distal stimulation from pups did not differentially activate Fos in any of 20 sites, including olfactory-processing structures such as the piriform cortex and medial amygdala. Physical interaction with pups, with or without suckling, elicited higher levels of Fos-immunoreactive nuclei than that of other conditions in numerous sites, including many previously implicated in maternal behaviour (medial preoptic nucleus, nucleus accumbens, lateral septum, lateral habenula, and the bed nucleus of the stria terminalis). Similar group patterns of Fos expression also occurred in sites not previously implicated in maternal behaviour (somatosensory cortex and paraventricular thalamic nucleus). Interaction with nonsuckling pups elicited the highest levels of Fos in the cortical amygdala, whereas suckling did not activate higher Fos than nonsuckling interaction in any site included in this report, including hypothalamic nuclei involved in lactation (paraventricular, supraoptic, and arcuate). There was little or no Fos in cingulate cortex, olfactory tubercle, medial septum, medial habenula, or ventromedial hypothalamus. These data suggest that trigeminal stimuli received by lactating rats during the performance of pronurturant maternal behaviour promote cellular activity resulting in neuronal expression of c-fos in many forebrain sites including the medial preoptic nucleus, several sites connected with it that are part of the mesotelencephalic dopamine system, and in the somatosensory cortex. In contrast, in these forebrain sites suckling does not elicit greater levels of Fos than that seen in nonsuckled rats and distal stimuli from pups are ineffective in increasing Fos levels compared with non-stimulated controls.

Effects of pH on the kinetic reaction mechanism of myoglobin unfolding studied by time-resolved electrospray ionization mass spectrometry.

In most cases, kinetic unfolding reactions of proteins follow a simple one-step mechanism that does not involve any detectable intermediates. One example for a more complicated unfolding reaction is the acid-induced denaturation of holo-myoglobin (hMb). This reaction proceeds through a transient intermediate and can be described by a sequential two-step mechanism (Konermann et al. Biochemistry 1997, 36, 6448-6454). Time-resolved electrospray ionization mass spectrometry (ESI MS) is a new technique for monitoring the kinetics of protein folding and unfolding in solution. Different protein conformations can be distinguished by the different charge state distributions that they generate during ESI. At the same time this technique allows monitoring the loss or binding of noncovalent protein ligands. In this work, time-resolved ESI MS is used to study the dependence of the kinetic unfolding mechanism of hMb on the specific solvent conditions used in the experiment. It is shown that hMb unfolds through a short-lived intermediate only at acidic pH. Under basic conditions no intermediate is observed. These findings are confirmed by the results of optical stopped-flow absorption experiments. This appears to be the first time that a dependence of the kinetic mechanism for protein unfolding on external conditions such as pH has been observed.

Functions of the caudal periaqueductal gray in lactating rats: kyphosis, lordosis, maternal aggression, and fearfulness.

Severe impairment of the kyphotic nursing posture in lactating rats found previously after prepartum lesions of the caudal intercollicular periaqueductal gray (cPAG-x) was confirmed and was extended to a continuous 24-hr period. Litters of cPAG-x dams gained approximately 10% less weight postnatally than controls, which was in part related to their dams' compensatory prone nursing posture that was ineffective for milk letdown. Sexual proceptivity and receptivity (lordosis) during the postpartum estrus were virtually eliminated in subjects with relatively large bilateral cPAG lesions. The doubling of maternal attacks toward a male intruder after lesioning was also confirmed and was related to reduced fearfulness in an elevated plus-maze. Thus, the cPAG plays a multifaceted role in parturient rats; it is involved in the mediation of nursing, sexual, aggressive, and fear behaviors.

Subungual tumors in incontinentia pigmenti.

A 22-year-old woman presented with intermittently painful subungual keratotic tumors of the hands as a late manifestation of incontinentia pigmenti. Scalloped bony deformities of the distal phalanges of both hands were noted on roentgenograms. The origin of these lytic deformities is uncertain; they may be secondary pressure phenomena or, alternatively, they may be an intrinsic manifestation of incontinentia pigmenti. These subungual tumors clinically resemble verrucae, keratoacanthomas, squamous cell carcinomas, or subungual fibromas. Therefore, it is important to recognize that subungual keratotic tumors may be a late manifestation of incontinentia pigmenti.

Hospital management of a patient with factitial dermatitis.

A 64-year-old male with a 40-year history of dermatologic disorders is presented. The patient carries diagnoses of neurodermatitis and suspected myosis fungoides on which has been superimposed a factitial dermatitis requiring increasingly extended hospitalizations. Given a very disturbed home life and personality style, the patient showed no incentive for change, and his family and caregivers became increasingly frustrated and distraught. An operant behavioral approach proved helpful to staff in standardizing care and setting realistic expectations. This led to a reversal of a downward course and to a coordinated discharge.

Overnight accommodations for visitors and outpatients: a nationwide study.

With the trend toward greater emphasis on ambulatory care and preadmission testing and the need to minimize travel and lodging costs for visitors and outpatients, a closer look at the hospital's role in arranging overnight accommodations becomes necessary. Unfortunately, little literature is available concerning the different types of overnight accommodations arranged by hospitals. The hospital interested in information on how it might upgrade its services to visitors and outpatients must rely primarily on brief descriptions of individual situations such as hospital-owned hotels or hostels and Ronald McDonald Houses. Although the literature indicates that some hospitals do offer some types of overnight accommodation services, the array of arrangements available and the level of hospital involvement is not always clear. Given this lack of information, the staff of the Patient and Visitor Participation Project of the University of Michigan Hospitals decided to conduct a nationwide study.

A forebrain-retrorubral pathway involved in male sex behavior is GABAergic and activated with mating in gerbils.

The ventral bed nuclei of the stria terminalis (BST) and medial preoptic nucleus (MPN) of gerbils contain cells that regulate male sex behavior via a largely uncrossed pathway to the retrorubral field (RRF). Our goal was to learn more about cells at the pathway source and target. To determine if the pathway uses GABA as its transmitter, we used immunocytochemistry (ICC) to study glutamic acid decarboxlyase(67) (GAD(67)) colocalization with fluoro-gold (FG) in the ventral BST and MPN after applying FG to the RRF. To determine if the pathway is activated with mating, we studied FG-Fos colocalization in the ventral BST of recently mated males. The ventral BST expresses Fos with mating and is the major pathway source. To determine to what extent other GABAergic cells in the ventral BST are activated with mating, we studied Fos colocalization with GAD(67) mRNA visualized by in situ hybridization (ISH). We also looked for GAD(67) mRNA in RRF cells. Almost all ventral BST and MPNm cells projecting to the RRF (95-97%) and most ventral BST cells activated with mating (89%), were GABAergic. GABAergic cells were also seen in the RRF. RRF-projecting cells represented 37% of ventral BST cells activated with mating. Their activation may reflect arousal and anticipation of sexual reward. Among ventral BST cells that project to the RRF, 14% were activated with mating, consistent with how much of this pathway is needed for mating. The activated GABAergic cells that do not project to the RRF may release GABA locally and inhibit ejaculation.

Functional interaction between the basolateral amygdala and the nucleus accumbens underlies incentive motivation for food reward on a fixed ratio schedule.

The ability for incentive properties of reward stimuli to maintain motivated behavior in the absence of the rewards themselves may be reliant in part on a glutamatergic projection from the basolateral (BLA) amygdala to the nucleus accumbens septi (NAS). The present work examined this idea in regard to food reward. In the first part of this study, lever pressing by rats on a fixed ratio 16 (FR16) schedule of food reinforcement was suppressed in a dose-dependent manner following bilateral infusion of the GABA(A) agonist muscimol to the BLA. Consumption of food when freely available was unaffected by the highest dose of muscimol, suggesting no change in the primary reward value of the food. Bilateral infusion of the broad-spectrum dopamine (DA) receptor antagonist flupenthixol to the NAS also resulted in a significant decrease in FR16 performance. As with the amygdala, consumption of freely available food was not affected by flupenthixol injections into the NAS. When unilateral injection of flupenthixol to the NAS was combined with contralateral injection of muscimol to the BLA, FR16 performance was suppressed. No significant change in lever press performance was observed following unilateral NAS injection of flupenthixol combined with ipsilateral injection of muscimol to the BLA. The results of this study support the idea that a functional connection between the BLA and NAS transmits incentive information necessary for the maintenance of responding in the absence of primary reward.

The immunochemistry of Shigella flexneri lipopolysaccharides. A quantitative analysis of their monosaccharide constituents.

1. The lipopolysaccharides of a representative selection of Shigella flexneri serotypes all contain the same constituents as Salmonella chemotype VII, namely, aldoheptose phosphate, 3-deoxy-2-oxo-octonate, O-phosphorylethanolamine, d-galactose, d-glucose, N-acetyl-d-glucosamine and l-rhamnose. 2. The presence of all the Salmonella basal sugars in Sh. flexneri lipopolysaccharides is consistent with the view that the latter contain a basal structure or core which is similar to the common basal structure of Salmonella lipopolysaccharides. 3. Although the Sh. flexneri lipopolysaccharides belong to one chemotype, there appear to be quantitative differences in the composition of their O-specific side chains. The repeating units of Sh. flexneri serotypes 1a, 2a, 3a, 4a, and variant X contain d-glucose, N-acetyl-d-glucosamine and l-rhamnose in the proportions 1:1:2 respectively. The analogous repeating units of serotypes 5a and 6 contain an additional mole of d-glucose and d-galactose respectively and that of variant Y 1 mole of d-glucose less.

Insulin does not regulate vascular smooth muscle Na+, K(+)-ATPase activity in rabbit aorta.

To determine whether insulin regulates vascular smooth muscle Na+, K(+)-ATPase activity and if impaired insulin stimulation of vascular smooth muscle Na+, K(+)-ATPase activity could be a cause of increased vascular reactivity to norepinephrine and angiotensin II in diabetic states, the effects of insulin on Na+, K(+)-ATPase activity were examined in normal rabbit aortic intima-media incubated with normal plasma glucose and myo-inositol levels for 30 min. Insulin at 100 microU/ml (600 pmol/l) had no effect on Na+, K(+)-ATPase activity. At 250 microU/ml it caused a 4.2 +/- 0.8% increase, and at 500 microU/ml insulin caused a 17.7 +/- 1.4% increase in Na+, K(+)-ATPase activity that was completely inhibited by amiloride (1 mmol/l). Human insulin-like growth factor I (600 pmol/l) caused an 18.0 +/- 1.0% increase in Na+, K(+)-ATPase activity that was inhibited by amiloride. Insulin does not regulate (stimulate) aortic vascular smooth muscle Na+, K(+)-ATPase activity. Supraphysiological insulin concentrations, probably acting through an insulin-like growth factor I receptor, stimulate Na+/H+ exchange in aortic vascular smooth muscle and cause small secondary increases in Na+, K(+)-ATPase activity. In aortic intima-media incubated with normal plasma glucose and myo-inositol levels, endogenously released adenosine stimulates and maintains a component of resting Na+, K(+)-ATPase activity and stimulates acute increases in activity when norepinephrine (1 mumol/l) or angiotensin II (100 nmol/l) is added. These adenosine-stimulated components of Na+, K(+)-ATPase activity are selectively inhibited when the medium glucose is raised to 30 mmol/l during a 30-min equilibration and 30-min incubation.(ABSTRACT TRUNCATED AT 250 WORDS)

Mechanism of glucose-induced (Na+, K+)-ATPase inhibition in aortic wall of rabbits.

Hyperglycaemia decreases (Na+, K+)-ATPase activity in specific tissues by a mechanism whose effects are prevented by aldose reductase inhibitors and by raising plasma myo-inositol. This mechanism was activated and studied in vitro in normal rabbit aortic intima-media. Raising medium glucose to 10 mmol/l for 60 min inhibited a major component of (Na+, K+)-ATPase-mediated 86Rb+/K+ uptake normally operative in resting aortic intima-media in medium containing normal plasma levels of glucose (5 mmol/l) and myo-inositol (70 mumol/l); 20 or 30 mmol/l glucose had no greater effect. This effect occurred under conditions in which the aortic intima-media's normal myo-inositol content is not detectably decreased. The inhibition was prevented by sorbinil (10 mumol/l) and by raising medium myo-inositol from 70 to 500 mumol/l, which had no effect on (Na+, K+)-ATPase activity when the medium glucose remained at 5 mmol/l. Raising medium glucose selectively inhibited a component of (Na+, K+)-ATPase activity that requires medium myo-inositol, because it is maintained by a regulatory system through rapid basal phosphatidylinositol turnover in a discrete pool, which is replenished by a fraction of basal de novo phosphatidylinositol synthesis that is selectively dependent on myo-inositol uptake. Medium myo-inositol at a normal plasma level became inadequate to maintain this fraction of basal de novo phosphatidylinositol synthesis [( 1,3-14C]glycerol incorporation) when the medium glucose was raised. When sorbinil was added raising medium glucose did not alter the ability of 70 mumol/l medium myoinositol to maintain the (Na+, K+)-ATPase activity that requires medium myo-inositol.(ABSTRACT TRUNCATED AT 250 WORDS)