Modulation by food restriction of intracellular calcium signaling in parotid acinar cells of aging Fischer 344 rats.

Previous studies suggest that alpha 1-adrenergic (alpha 1-AR)-induced intracellular calcium ([Ca2+]i) mobilization in rat parotid acinar cells declines with age. In this study, we examined the effects of food restriction on alpha 1-AR and muscarinic-stimulated [Ca2+]i mobilization in parotid acinar cells during aging. [Ca2+]i levels in response to the alpha 1-AR agonist epinephrine and the muscarinic agonist carbachol were evaluated in Fura-2-loaded parotid acinar cells from ad libitum-fed (AL) and food-restricted (FR) Fischer 344 male rats at 4, 6, 14, and 24 months of age. [Ca2+]i responses to epinephrine and carbachol (10 microM) were significantly reduced (48% and 35%, respectively; p < .05) in cells from 24-month-old AL rats as compared to younger AL rats. In contrast, no significant reduction of epinephrine and carbachol responses was observed in 24-month-old FR animals. An age-related increase in basal [Ca2+]i (peak around 14 months; p < .02) was observed in both AL and FR rats. In addition, basal [Ca2+]i was higher in FR than in AL rats at 14 and 24 months of age (p < .02). These studies suggest that FR partially attenuates or delays age-related impairments in alpha 1-AR- and muscarinic-cholinergic signal transduction systems of parotid acinar cells. Basal [Ca2+]i also appears to be altered during aging and by FR.

Efficiency of lower threshold criteria for the diagnosis of gestational diabetes.

OBJECTIVES: To determine the incidence of adverse outcome in normal untreated gravidas with minimal hyperglycemia, classified as having gestational diabetes mellitus (GDM) by threshold criteria lower than current standards; to determine how efficient the different criteria are in identifying infants at risk for morbidity; and to explore the pathophysiology of minimal hyperglycemia using the glucose tolerance test (GTT) periodicity concept. METHODS: Seven hundred eight subjects considered nondiabetic by current ACOG criteria were reclassified by the criteria of Coustan (fasting 95, 1 hour 180, 2 hours 155, and 5 hours 140 mg/dL), Sacks (96, 172, 152, and 131 mg/dL), or Langer (at least one abnormal ACOG value). Glucose tolerance test periodicity, the incidence of large for gestational age (LGA) neonates, and macrosomia were then determined for each gravida diagnosed as having GDM by these criteria. RESULTS: Both Coustan and Langer criteria identified a significantly greater incidence of LGA infants compared with non-GDM (23.6 and 25.3%, respectively, versus 14%, P < .05), and identified them as efficiently as current criteria, approximately one LGA infant for every four GDM subjects treated. The incidence of LGA did not differ between the Sacks GDM and non-GDM groups. Glucose tolerance test periodicity for newly diagnosed GDM gravidas was significantly longer than non-GDM for Coustan and Langer criteria (3.9 and 4.06 versus 3.3 hours, P < .01) but not for the Sacks criteria. CONCLUSION: Using lower threshold criteria to diagnose GDM identified morbidity at an incidence and efficiency comparable to current standards. These newly diagnosed GDM gravidas had abnormal GTT characteristics, with each group exceeding the 3.5-hour GTT periodicity limit previously found for nondiabetic gravidas. Sack's conversion of existing standards may be too low to efficiently identify pregnant subjects at risk for increased morbidity.

Treatment of murine cryptosporidiosis with anticryptosporidial immune rat bile.

Persistent cryptosporidiosis was established in nu/nu BALB/c mice by oral inoculation with Cryptosporidium parvum oocysts. The model was used to determine the impact of anticryptosporidial immune rat bile on the resolution of the disease. Presence of C. parvum-specific IgA in the immune rat bile was determined by enzyme-linked immunosorbent assay. Infection of mice was verified by stool analysis for oocytes and by hematoxylin and eosin-stained intestinal sections from control mice (infected but untreated). Efficacy of treatment was determined in control and treated mice by analysis of identical, hematoxylin and eosin-stained sections of the small intestine and cecum. Semi-quantitative comparisons were made by determining the percent of crypts infected with Cryptosporidium organisms. The scores of treated mice were significantly lower then controls. Microscopic analysis of intestinal sections showed less villus atrophy, crypt hyperplasia, and fewer organisms per crypt in the immune bile-treated mice than in controls. These results support a role for humoral immunity in the eradication of cryptosporidiosis.

Cocaine impairs follicular phase pulsatile gonadotropin secretion in rhesus monkeys.

OBJECTIVE: To assess cocaine's effect on follicular phase pulsatile gonadotropin secretion in normally cycling rhesus monkeys. METHODS: Sixteen monkeys were paired by body weight and randomized to receive intravenous saline (n = 8) or cocaine (4 mg/kg, n = 8) daily on cycle days 2 to 14. Monkeys were chronically cannulated to allow frequent blood collections without anesthesia. Blood samples were obtained every 15 minutes for 8 hours in early (EFP; cycle days 1 to 5), mid-(MFP; cycle days 6 to 10), and late (LFP; cycle days 11 to 15) follicular phase. Plasma concentrations of LH, FSH, and estradiol-17 beta (E2) were determined by radioimmunoassay. Pulses were identified by cluster analysis. Statistical differences were determined by analysis of variance (ANOVA) and Sidak's multiple comparison test. RESULTS: Seven out of eight monkeys in the control group demonstrated timely ovulation. Only one monkey in the cocaine-treated group ovulated. Similar gonadotropin pulse intervals (70 to 90 minutes) were observed throughout the follicular phase in both the controls and cocaine-treated monkeys. LH and FSH pulse amplitudes increased significantly from the EFP/MFP to the LFP in controls. In cocaine-treated monkeys, gonadotropin pulse amplitudes remained at EFP/MFP levels throughout the study period. The mean gonadotropin pulse amplitude and the mean E2 levels in the LFP were significantly greater in controls as compared with cocaine-treated monkeys (P < .001). CONCLUSION: These findings demonstrate that cocaine suppresses the normal increase in LH and FSH pulse amplitude seen in the LFP. Further studies are in progress to determine the mechanism of cocaine's disruption of the hypothalamic-pituitary-ovarian axis.

Low-dose follicular-phase cocaine administration disrupts menstrual and ovarian cyclicity in rhesus monkeys.

OBJECTIVE: To evaluate the effects of daily low-dose follicular-phase cocaine administration on menstrual cyclicity, ovulation rates, corpus luteum function, and hormone levels in rhesus monkeys. METHOD: Normally cycling, drug-naive, adult rhesus monkeys were randomized to receive either 1 mg/kg of cocaine (n = 7), 2 mg/kg of cocaine (n = 7), or normal saline (n = 7) daily on cycle days 2 to 14. Daily blood samples were obtained through indwelling catheters for measurement of serum gonadotropins and ovarian steroids. Daily vaginal swabs were obtained to determine onset of menses. Laparoscopy was performed 2 days after the midcycle estrogen peak to document ovulation. Daily caloric intakes as well as pretreatment and posttreatment weights were recorded. RESULTS: Two of seven monkeys receiving 1 mg/kg per day and two of seven monkeys receiving 2 mg/kg per day of cocaine had timely ovulation and normal menstrual cycle lengths. One monkey receiving the 2-mg/kg dose ovulated on cycle day 24 and had a short luteal phase (7 days) with a mean progesterone level of 2.4 ng/mL. All seven saline-treated control monkeys ovulated normally; the mean cycle length was 29 days and all had adequate luteal phases. The difference in ovulation rates between cocaine-treated and control monkeys was statistically significant (P = .003). There were no differences in basal levels of LH or FSH between treatment groups. There were no significant differences in weight change or caloric intake among groups. One third of the subsequent menstrual cycles in cocaine-treated monkeys were of abnormal duration. CONCLUSION: Daily low-dose follicular-phase cocaine administration disrupts menstrual cyclicity and folliculogenesis. This effect is independent of weight loss, caloric intake, and basal gonadotropin levels. Cocaine exposure may have a persistent effect on menstrual and ovarian cyclicity in some monkeys.

Treatment of experimental systemic mycoses with BRL 49594A.

BRL 49594A (BRL) is a water soluble analogue of amphotericin B which has been developed as a polyene with efficacy similar to amphotericin B but with much reduced toxicity. BRL was prepared in 5% glucose, and was used to treat mice experimentally infected with Aspergillus fumigatus, Cryptococcus neoformans, or Histoplasma capsulatum. In the models in which BRL and amphotericin B were compared, BRL was well tolerated but was less effective than a similar regimen of amphotericin B. However, the ability to give much larger doses of BRL than tolerated with amphotericin B suggest that this drug could be an alternative to amphotericin B.

Treatment of murine cryptococcal meningitis with UR-9751 and UR-9746.

In these studies, we compare the efficacy of two new azole antifungals with fluconazole in a murine model of cryptococcal meningitis. Mice were infected intracranially. Beginning one day later, groups of 7-10 mice were treated through to day 10 orally with UR-9751 or UR-9746 at 0.1, 0.25, 0.5, 1 or 10 mg kg(-1) day(-1) or fluconazole at 10 mg kg(-1) day(-1). At 10 mg kg(-1) day(-1), all three drugs prolonged survival over controls, but at 1 mg kg(-1) day(-1) only UR-9746 prolonged survival. Tissue counts were more varied on mice sacrificed 8 days after infection. In general, both UR drugs were equal or more potent than fluconazole, and UR-9751 was more effective than UR-9746.

Treatment of murine disseminated candidiasis with L-743,872.

L-743,872 (M991), which is a pneumocandin derivative, was evaluated in a mouse model of disseminated candidiasis caused by a fluconazole-resistant isolate of Candida albicans. In immunocompetent mice M991 prolonged survival at doses as low as 0.0125 mg/kg of body weight per day. In neutropenic mice 0.05 mg/kg was the lowest effective dose. M991 is a very potent drug for treatment of disseminated candidiasis.

Stroke in infective endocarditis.

We reviewed 212 consecutive episodes of infective endocarditis in 203 patients at six hospitals between 1978 and 1986 and found that 21% were complicated by stroke. Of 133 episodes involving native mitral and/or aortic valves, brain ischemia occurred in 19%, brain hemorrhage in 7%, and non-central nervous system emboli in 11%; vegetations were identified in 56% of 113 adequate echocardiograms and did not correlate with risk of embolism. In native-valve endocarditis, most (74%) ischemic strokes had occurred by the time of presentation and an additional 13% occurred less than or equal to 48 hours after diagnosis; the incidence of brain ischemia was 13% on presentation, 3% during the first 48 hours of hospitalization, and 2%-5% during the remainder of the acute course. Stroke recurred at a rate of 0.5%/day, often heralding relapse/uncontrolled infection. Only 9% of ischemic infarcts were large (all in patients with Staphylococcus aureus infection), while 8% were small and subcortical. Brain hemorrhage occurred primarily at the time of presentation, particularly in intravenous drug abusers, and was associated with uncontrolled S. aureus infection with pyogenic arteritis. Ischemic and hemorrhagic stroke continue to be frequent and important in patients with infective endocarditis and are clustered during uncontrolled infection.(ABSTRACT TRUNCATED AT 250 WORDS)

ZD0870 treatment of murine candidiasis caused by fluconazole resistant isolates of Candida albicans.

Mice were infected intravenously with three fluconazole susceptible and ten fluconazole resistant isolates of Candida albicans then treated with escalating doses of 0.25, 0.5, 1, 2.5, 10 and 40 mg/kg/day of the new antifungal triazole, ZD0870, for 10 days. A minimum protective dose of < 0.25 mg/kg was determined for infections introduced by the three fluconazole susceptible C. albicans and one of the fluconazole resistant isolates whereas doses ranging from 2.5 to 10 mg/kg/day were required for infections induced by seven of the resistant isolates and > or = 40 mg/kg/day for the remainder. Thus, infections caused by fluconazole resistant C. albicans may be successfully treated with ZD0870, though higher doses than those used to treat infections due to susceptible yeast may be required.

Granulocyte colony stimulating factor therapy of experimental cryptococcal meningitis.

Cryptococcal meningitis was induced in mice using intracerebral injection of Cryptococcus neoformans. Beginning either 3 days before or 1 day after infection, mice were treated with human recombinant granulocyte colony stimulating factor (hGCSF). In high doses hGCSF reduced the brain tissue burden of C. neoformans but had no effect on survival. The effect of hGCSF was dependent on size of the infecting dose and time of administration. A large innocula of C. neoformans, or when hGCSF was initiated after infection, there was no added benefit. Some groups of mice also received low doses of fluconazole beginning 1 day after infection. Fluconazole both prolonged survival and reduced brain tissue counts of C. neoformans. Combined cytokine/fluconazole therapy was superior to either agent given alone. These studies suggest that hGCSF can add to the efficacy of fluconazole therapy in murine cryptococcosis, and suggest that polymorphonuclear leucocytes contribute to host defence in cryptococcal meningitis. The relative potency of fluconazole appears greater than hGCSF.

Granulocyte colony-stimulating factor and azole antifungal therapy in murine aspergillosis: role of immune suppression.

Outbred ICR mice were immune suppressed either with hydrocortisone or with 5-fluorouracil and were infected intranasally with Aspergillus fumigatus. Beginning 3 days before infection some groups of mice were given recombinant human granulocyte colony-stimulating factor (G-CSF), SCH56592 (an antifungal triazole), or both. Corticosteroid-pretreated mice responded to SCH56592 and had reduced counts in lung tissue and prolonged survival. In these mice, G-CSF strongly antagonized the antifungal activity of SCH56592. Animals treated with both agents developed large lung abscesses with polymorphonuclear leukocytes and large amounts of Aspergillus. In contrast, mice made neutropenic with 5-fluorouracil and then infected with A. fumigatus conidia benefited from either G-CSF or triazoles, and the effect of the combination was additive rather than antagonistic. Host predisposing factors contribute in different ways to the outcome of growth factor therapy in aspergillosis.

Fluconazole, D0870, and flucytosine treatment of disseminated Candida tropicalis infections in mice.

D0870 is a recently developed triazole with characteristics of a broad spectrum of activity and slow clearance by nonrenal mechanisms. Herein we have evaluated the efficacy of D0870, alone and combined with flucytosine, in a murine model of disseminated Candida tropicalis infection. Four isolates of C. tropicalis were evaluated. Two were highly susceptible in vitro to fluconazole, and two were resistant to fluconazole. All were highly susceptible to flucytosine and D0870. Animals were pretreated with 5-fluorouracil 1 day before infection because C. tropicalis has reduced virulence in immunocompetent mice. This was done to render them neutropenic for > 10 days. Mice were infected intravenously and treated orally with D0870 or fluconazole, alone or combined with flucytosine. Survival and tissue burden of the spleen and kidneys were used to evaluate the efficacy of antifungal therapy. Fluconazole was less effective for treatment of resistant C. tropicalis than susceptible C. tropicalis. D0870 was more potent than fluconazole and was effective in fluconazole-resistant isolates. Flucytosine was consistently effective when used alone but did not consistently add to the benefit of D0870 or fluconazole. D0870 has potential in treatment of candidiasis caused by C. tropicalis, including fluconazole-resistant isolates.

Efficacy of nikkomycin Z in the treatment of murine histoplasmosis.

Immune-competent ICR and BALB/c athymic (nude) mice were infected intravenously with Histoplasma capsulatum and treated with either fluconazole or nikkomycin Z or 5% dextrose (controls). In immune-competent ICR mice, fluconazole and nikkomycin Z both prolonged survival when given at 5 mg/kg of body weight twice daily. When administered in doses as low as 2.5 mg/kg twice daily, nikkomycin Z reduced fungal counts in both the spleen and liver. When both drugs were combined, there was no antagonism, and in combined therapy spleen and liver counts were reduced more than for either drug alone. However, nikkomycin Z had no effect on brain fungal burden. In nude mice fluconazole and nikkomycin Z had an additive effect in prolongation of survival and reduction of liver and spleen burden. Nikkomycin Z is well tolerated, is at least as effective as fluconazole, and may interact beneficially with fluconazole for treatment of murine histoplasmosis.

SCH56592 treatment of murine invasive aspergillosis.

Mice were immunosuppressed using corticosteroids and infected with conidia of Aspergillus fumigatus. Beginning 1 day after infection, mice were treated orally either with Noble agar or with SCH56592 suspended in Noble agar, or intraperitoneally with amphotericin B. SCH56592 prolonged survival and reduced lung tissue counts of A. fumigatus, while amphotericin B had marginal benefit. SCH56592 merits further development for treatment of aspergillosis.

Fluconazole treatment of Candida albicans infection in mice: does in vitro susceptibility predict in vivo response?

A series of fluconazole-susceptible and-fluconazole resistant Candida albicans fungal isolates were used to infect mice intravenously. Mice were treated with varying doses of fluconazole beginning one day after infection. For all of the 6 fluconazole-susceptible isolates, fluconazole was highly effective at <0.25 mg/kg of body weight twice daily. By contrast, fluconazole was less effective in at least 6 of 10 fluconazole-resistant isolates and was ineffective at > or = 40 mg/kg twice daily in 4 fluconazole-resistant isolates. Although the correlation is not precise, in vitro susceptibility testing of C. albicans can predict in vivo response to fluconazole.

Cocaine impairs gonadotropin secretion in oophorectomized monkeys.

OBJECTIVE: Our objective was to determine whether cocaine alters gonadotropin secretion in oophorectomized monkeys. STUDY DESIGN: Oophorectomized monkeys with elevated gonadotropin levels were chronically cannulated to allow blood sampling every 15 minutes. Monkeys received either saline solution or 2 or 4 mg/kg cocaine hydrochloride as an intravenous bolus. Other oophorectomized monkeys were pretreated with either saline solution or 4 mg/kg cocaine 2 hours before bolus gonadotropin-releasing hormone administration, and plasma luteinizing hormone and follicle-stimulating hormone levels were measured every 15 minutes for 3 hours. Monkeys were also given either saline solution or 4 mg/kg of cocaine with gonadotropin-releasing hormone simultaneously, and plasma gonadotropin levels were measured every 15 minutes for 3 hours. Serum luteinizing hormone and follicle-stimulating hormone levels were measured by radioimmunoassay. RESULTS: Both doses of cocaine resulted in a significant decrease in luteinizing hormone levels compared with controls. Follicle-stimulating hormone levels were significantly decreased only with the 4 mg/kg dose of cocaine. There was no difference in luteinizing hormone and follicle-stimulating hormone responses to gonadotropin-releasing hormone in the cocaine-treated monkeys compared with saline solution-treated monkeys by using repeated-measures analysis of variance. CONCLUSION: These findings demonstrate that acute cocaine administration to oophorectomized primates inhibits basal luteinizing hormone-follicle-stimulating hormone secretion but not gonadotropin-releasing hormone-stimulated luteinizing hormone and follicle-stimulating hormone release. In the absence of an effect on gonadotropin-releasing hormone-stimulated gonadotropin release, we conclude that the impaired luteinizing hormone-follicle-stimulating hormone secretion after cocaine administration is due in part to a direct effect of cocaine on gonadotropin-releasing hormone neurons or on hypothalamic neurotransmitter modulation of gonadotropin-releasing hormone release.

Treatment of histoplasmosis with MK-991 (L-743,872).

BALB/c nu/+ immunocompetent and athymic (nu/nu) mice were infected intravenously with yeast cells of Histoplasma capsulatum. Mice were either given water (controls) intraperitoneally (i.p.) or given MK-991 i.p. once daily or twice daily. Protection was measured as prolonged survival or reduction in tissue counts. MK-991 was protective in immunocompetent mice, prolonging survival and reducing counts in spleen and livers at a dose as low as 0.05 mg/kg of body weight/day. MK-991 was modestly effective in athymic mice at a higher dose, 5 mg/kg/day. These studies suggest that MK-991 may be appropriate for clinical development in histoplasmosis.

Variation in fluconazole efficacy for Candida albicans strains sequentially isolated from oral cavities of patients with AIDS in an experimental murine candidiasis model.

Four strains of Candida albicans, isolated from two patients with AIDS who had undergone prolonged fluconazole therapy for oral candidiasis, were studied in a model of disseminated murine candidiasis. Pre- and posttreatment isolates from each patient were genetically related, and the fluconazole MICs for the strains had increased significantly, from 0.25 to 32 micrograms/ml for the strains isolated from patient 1 and from 1.0 to 16 micrograms/ml for the strains isolated from patient 2. Mice were infected intravenously and were treated orally with fluconazole. For survival studies, mice were treated from day 1 to day 10 postinfection and were observed through day 30. The fluconazole dosages were as follows: 0.25, 0.5, 1.0, and 5.0 mg/kg of body weight twice a day. For tissue burden studies, two groups of mice (each group received fluconazole at 0.25 or 5.0 mg/kg) were treated from day 1 to day 7 and were sacrificed 1 day later for quantitative tissue cultures of the spleen and both kidneys. For pretreatment isolates from both patients, all fluconazole dosing regimens were effective at prolonging survival compared with the survival of the control groups. For posttreatment isolates, only fluconazole at 5.0 mg/kg was effective at prolonging survival. Both fluconazole dosing regimens used in the tissue burden studies significantly reduced the counts of the pretreatment isolate from patient 1 in the spleen and kidney, while fluconazole at 5.0 mg/kg was effective at reducing the counts of the posttreatment isolate. For both isolates from patient 2, only fluconazole at 5.0 mg/kg was effective at reducing the counts in the spleen and kidney. The study indicates that C. albicans mutation to resistance to fluconazole may play a critical role in fluconazole-refractory oral candidiasis in AIDS patients.

Comparison of fluconazole, amphotericin B and flucytosine in treatment of a murine model of disseminated infection with Candida glabrata in immunocompromised mice.

Candida glabrata is an emerging opportunist pathogen in immunosuppressed patients. C. glabrata is resistant to many antifungal agents and until recently, there have been no standard treatment regimens for this organism. A mouse model was established using mice immunosuppressed with 5 fluorouracil to evaluate amphotericin B, flucytosine, fluconazole and their combinations to treat an intravenously induced C. glabrata infection. Treatment with fluconazole, flucytosine, amphotericin B or a combination was begun one day after infection. Following 5 days of treatment, the mice were killed for fungal counts in kidneys and spleen. At the doses used, amphotericin B was superior to fluconazole or flucytosine alone in the treatment of C. glabrata infections. Flucytosine reduced the fungal burden in the kidney for only two of four isolates of C. glabrata. The combination of fluconazole and flucytosine was superior to these agents alone in reducing the tissue burden in the kidney for one isolate of C. glabrata. High doses of fluconazole alone produced modest reductions in kidney counts but did not reduce spleen tissue counts. There was poor correlation between in-vitro MICs and in-vivo results.