Relaxin and decidualization in mice: a reappraisal

The nature of the physiological stimulus inducing decidualization in the endometrium is unknown. In this study we attempted to verify a recent report that relaxin can induce decidualization in intact mice primed with a high dose of estradiol valerate (5 micrograms) and a low dose (10 micrograms) of medroxyprogesterone acetate. In our study, neither s.c. nor intrauterine relaxin, nor intraluminal arachis oil, (an established deciduogenic stimulus) were able to induce decidualization. In addition, while oil was able to induce decidualization (increased uterine weight, and positive Pontamine Sky Blue and stromal alkaline phosphatase reactions) in ovariectomized mice treated with a regimen of estradiol and medroxyprogesterone acetate designed to produce optimum uterine sensitivity, no decidualization occurred in response to either s.c. or intraluminal relaxin. This study fails to provide any support for a role for relaxin as a deciduogenic stimulus.     

Toxicokinetics of isoprene in rodents and humans

A physiological toxicokinetic model (PT model) was developed for inhaled isoprene in mouse, rat and man. Partition coefficients blood:air and tissue:blood were determined in vitro by a headspace method. Parameters of a saturable isoprene metabolism in B6C3F1 mice, Sprague-Dawley rats and volunteers were obtained from gas uptake experiments in closed systems, analyzed by means of a two-compartment model. Incorporation of these parameters into the PT model revealed that isoprene was metabolized not only in the liver but also in extrahepatic organs. Endogenous production of isoprene in man was quantified from experiments with volunteers breathing into a closed system. The PT model was validated for mice, rats and humans by comparing simulated values with data determined by other authors.     

Dexamethasone-induced changes in endometrial growth and inducible nitric oxide synthase during decidualization in rats

1. The present study investigated the time-dependent inhibitory responses of endometrial growth and inducible nitric oxide synthase (iNOS) to dexamethasone during deciduoma development that was surgically induced on day 4 of pseudopregnancy (PG). 2. Groups of rats (n = 6) were subcutaneously injected with dexamethasone (1.5 mg/rat per day) for 3 days (PG days 1-3, 4-6, 7-9, 10-12 and 12-15). Rats in each group were killed on the last injection day. 3. Dexamethasone produced comparable temporal inhibitory changes in endometrial growth (wet weight, protein and DNA concentrations; P<0.0001) and in iNOS activity (130 kDa protein band), which peaked after PG days 4-6 and 7-9 pretreatments. 4. Endometrial matrix metalloproteinases (72 and 92 kDa) activity profiles displayed maximal reductions (36 and 53%, respectively) following PG days 4-6 pretreatment. Serum progesterone levels were equally (P<0.0001) but asynchronously inhibited by dexamethasone on PG days 9 and 12. 5. Dexamethasone inhibition of endometrial growth and in situ iNOS was most pronounced during decidual development (PG days 4-9). Minor reductions in these endometrial parameters occurred before deciduoma induction (PG days 1-3) and during deciduoma regression (PG days 10-15). 6. These results indicate that, in the endometrium, the iNOS/endogenous nitric oxide system may be linked to the biochemical and metabolic mechanisms responsible for the developmental responsiveness of the deciduoma to dexamethasone exposure. These time-dependent changes in endometrial growth and iNOS apparently were not mediated by progesterone.     

Fetal islet xenotransplantation in rodents and primates

(E)-2'-deoxy-2'-(fluoromethylene) cytidine (FMdC), a novel inhibitor of ribonucleotide-diphosphate reductase, has been shown to have anti-tumor activity against solid tumors and sensitize tumor cells to ionizing radiation. Pentoxifylline (PTX) can potentiate the cell killing induced by DNA-damaging agents through abrogation of DNA-damage-dependent G2 checkpoint. We investigated the cytotoxic, radiosensitizing and cell-cycle effects of FMdC and PTX in a human colon-cancer cell line WiDr. PTX at 0.25-1.0 mM enhanced the cytotoxicity of FMdC and lowered the IC50 of FMdC from 79 +/- 0.1 to 31.2 +/- 2.1 nM, as determined by MTT assay. Using clonogenic assay, pre-irradiation exposure of exponentially growing WiDr cells to 30 nM FMdC for 48 hr or post-irradiation to 0.5 to 1.0 mM PTX alone resulted in an increase in radiation-induced cytotoxicity. Moreover, there was a significant change of the radiosensitization if both drugs were combined as compared with the effect of either drug alone. Cell-cycle analysis showed that treatment with nanomolar FMdC resulted in S-phase accumulation and that such an S-phase arrest can be abrogated by PTX. Treatment with FMdC prior to radiation increased post-irradiation-induced G2 arrest, and such G2 accumulation was also abrogated by PTX. These results suggest that pharmacological abrogation of S and G2 checkpoints by PTX may provide an effective strategy for enhancing the cytotoxic and radiosensitizing effects of FMdC.     

Requirement for transglutaminase in progesterone-induced decidualization of human endometrial stromal cells

Differentiation of endometrial stromal cells (decidualization) is essential for embryo implantation and maintenance of pregnancy. By sequential complementary DNA subtractive hybridization, one of the messenger RNAs (mRNA) induced by progesterone in human endometrial stromal cells decidualized in vitro was identified as that of a tissue transglutaminase type II (TGase). TGase mRNA was induced within 6 h after the addition of progesterone to the culture, and the effect was dose dependent. Both the TGase inhibitor monodansylcadaverine and oligodeoxynucleotide complementary to the TGase mRNA inhibited the decidualization, as assessed by PRL production and morphological transformation. Expression of TGase mRNA in human decidua and endometria exposed to high levels of progesterone in vivo was demonstrated by Northern blotting and in situ hybridization. These data suggest that TGase is necessary for the decidualization of human endometrial stromal cells and that clarification of the mechanism of action of TGase will facilitate further insight into the diagnosis and treatment of infertility.     

Ketamine attenuates and reverses morphine tolerance in rodents

BACKGROUND: The development of tolerance complicates the use of morphine to manage persistent pain. N-methyl-D-aspartate receptor antagonists can attenuate or reverse morphine tolerance. The authors studied ketamine's ability to modulate morphine tolerance. METHOD: Tolerance was produced in mice given morphine subcutaneously and was assessed by a cumulative dose-response analysis using the tail-flick test. The ability of ketamine at 0.3, 3, or 10 mg/kg given subcutaneously before and after morphine to attenuate the development of tolerance was assessed. The ability of 10 mg/kg ketamine to reverse tolerance produced by the subcutaneous implantation of morphine pellets to mice was also assessed. Rats were made tolerant to intraspinal morphine and the effects of the coadministration of 12 micrograms intraspinal ketamine were assessed. RESULTS: Morphine given subcutaneously produced a fivefold increase in the median effective (ED50) dose of morphine, which was dose-dependently attenuated by subcutaneously administered ketamine. A tenfold increase in the morphine ED50 produced by morphine pellets was completely reversed by ketamine given subcutaneously. Intraspinal morphine produced a 46-fold increase in its ED50, which was almost completely attenuated by the coadministration of intraspinal ketamine. CONCLUSIONS: Systemically administered ketamine attenuates and reverses systemically induced morphine tolerance in mice, and intraspinal ketamine attenuates tolerance produced by intraspinal morphine in rats.     

Progesterone-dependent decidualization of the human endometrium is mediated by cAMP

BACKGROUND: Behavioral stress has been proposed to contribute to the occurrence of myocardial ischemia. Objective To investigate the effect of chronic exposure to behavioral stress on the function and structure of the coronary artery of borderline hypertensive rats (BHR). DESIGN: BHR were either exposed to an air-jet stress for 2 h/day for 10 days or kept in their cage for 10 days. METHODS: After 10 days, hemodynamic measurements in conscious animals were recorded, and their hearts were removed for isolation of a left ventricular coronary artery for functional studies or for fixation by retrograde perfusion for study with scanning electron microscopy. Vascular reactivity was measured in isolated coronary arteries (approximately 250 microm) maintained at an intraluminal diameter of 40 mmHg while the intraluminal diameter was recorded continuously. RESULTS: The resting mean arterial pressure and heart rate in conscious, unrestrained BHR were not altered significantly by exposure to 10 days of 2 h/day air-jet stress. Coronary artery relaxation in response to the endothelium-dependent vasodilator acetylcholine was impaired in rats exposed to the air-jet stress compared with that in controls. An attenuated response to exogenous nitric oxide in coronary arteries from stressed BHR was confirmed by the finding of a reduced sensitivity to nitroprusside, which releases nitric oxide independently from the endothelium. However, relaxation of coronary arteries in response to isoproterenol, which acts independently from nitric oxide, was not altered. Coronary artery contraction in response to endothelin-1 and phenylephrine was not altered in vessels taken from BHR exposed to behavioral stress compared with that in vessels from control rats. Scanning electron microscopy of the endothelial surface of the septal coronary artery showed no difference between vessels from control and stressed BHR. CONCLUSION: These results indicate that behavioral stress impairs endothelium-dependent and nitric oxide-mediated coronary relaxation, but does not alter alpha1-adrenoceptor or endothelin-1-mediated contraction. By impairing coronary artery vascular relaxation, chronic exposure to behavioral stress may contribute to myocardial ischemia.     

Weak male-driven molecular evolution in rodents

In humans and rodents the male-to-female ratio of mutation rate (alpha m) has been suggested to be extremely large, so that the process of nucleotide substitution is almost completely male-driven. However, our sequence data from the last intron of the X chromosome-linked (Zfx) and Y chromosome-linked (Zfy) zinc finger protein genes suggest that alpha m is only approximately 2 in rodents with a 95% confidence interval from 1 to 3. Moreover, from published data on oogenesis and spermatogenesis we estimate the male-to-female ratio of the number of germ cell divisions per generation to be approximately 2 in rodents, confirming our estimate of alpha m and suggesting that errors in DNA replication are the primary source of mutation. As the estimated alpha m for rodents is only one-third of our previous estimate of approximately 6 for higher primates, there appear to be generation-time effects--i.e., alpha m decreases with decreasing generation time.     

Hemorrhagic gastritis in free-living rodents in Idaho

Between February 1992 and March 1994, four species of rodent from the Snake River Birds of Prey Area near Boise, Idaho (USA) were necropsied. Hemorrhagic gastritis was observed in 16 of 131 Townsend's ground squirrels (Spermophilus townsendii), one of 11 Ord's kangaroo rats (Dipodomys ordii) and the one Great Basin pocket mouse (Perognathus parvus) evaluated. No lesions were observed in 14 white-footed deer mice (Peromyscus maniculatus). Tissue from one Townsend's ground squirrel was negative for Helicobacter sp.-like bacteria.     

Regulation of ob gene expression in rodents and humans

The discovery of the obese gene in the mouse and its conserved homologue in humans has led to important discoveries in energy metabolism. One of the chief findings was the fact that the expression of the leptin gene was regulated and that it, in turn, could regulate metabolism and behavior. Much of the literature has focused on the physiological role of leptin in driving processes as diverse as reproduction, starvation defence, feeding behavior or body weight, all dependent on expression levels of the ob gene. Here, we will describe our work, in which we have begun to elucidate the regulatory processes controlling obese gene expression.     

Angiotensin III-induced dipsogenic and pressor responses in rodents.

Three experiments examined responses to angiotensins in 64 male Sprague-Dawley rats, 64 male Mongolian gerbils, and 40 Octodon degus—a South American rodent. In Exp I, injections of [des-Asp–1]-angiotensin I ([des-Asp–1]-AI), angiotensin II (AII), and angiotensin III (AIII), at doses of .001–2 mg/kg (sc), induced drinking in the rat and degus, but not in the gerbil. In Exp II, pretreatment with captopril (50 mg/kg), an angiotensin converting enzyme inhibitor, prevented the endogenous conversion of sc injected [des-Asp–1]-AI to AIII and prevented drinking in rats and degus. The pharmacological artifact of hypovolemia caused by angiotensin-induced increases in vascular permeability was not observed in members of these species. In Exp III, blood pressure changes resulting from injections of AII and AIII in rats and gerbils were measured. Significant pressor elevations were seen following the administration of both analogs, although AII was more potent. Results demonstrate that AIII is dipsogenic in rats and degus and serves as a pressor agent in rats and gerbils. No explanation was found for the gerbil's relative lack of dipsogenicity to the presently tested angiotensins. (54 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Hypolipidemic activity of boronated nucleosides and nucleotides in rodents

Base-boronated nucleoside and phosphate-boronated nucleotides were potent hypolipidemic agents in rodents, lowering both serum cholesterol and triglyceride levels. Rat VLDL and LDL cholesterol levels were generally reduced and HDL cholesterol levels were significantly elevated after 14 days dosing at 8 mg/kg/day. Tissue cholesterol, triglyceride and phospholipid levels were reduced by selected derivatives. Increased fecal excretion of lipids did not appear to be a mechanism by which these derivatives lowered serum lipids in rodents. Rather, the agents suppressed appetite and reduced the activities of rate-limiting enzymes for de novo lipid synthesis, specifically cytoplasmic acetyl CoA synthetase, squalene synthetase, and phosphatidylate phosphohydrolase with IC50 values of approximately 10(-5) m.     

Comparative studies on the in vitro decidualization process in the baboon (Papio anubis) and human

Blood supply is essential for the maintenance of epididymal function. Since there is no considerable neovascularization in the epididymis, this tissue could represent a suitable model to study the vascular endothelial growth factor (VEGF) effect for vascular permeability. We studied the expression and function of VEGF and its receptors fms-like tyrosine kinase (Flt-1) and fetal liver kinase (designated as kinase insert domain-containing receptor, KDR in the human) in the human epididymis. VEGF and VEGF receptors mRNA were detected in the human epididymal tissue. VEGF protein was localized in peritubular and in ciliated cells of efferent ducts as well as in peritubular and basal cells of the epididymal duct. Vascular endothelial cells did not express VEGF. Flt-1 protein was localized in ciliated cells of efferent ducts and in lymphatic vessels. Vascular endothelial cells were negative for Flt-1 but positive for KDR. In vitro VEGF165 treatment of epididymal tissue induced endothelial fenestrations and opening of interendothelial junctions. Additionally, we observed for the first time that VEGF could induce transendothelial gaps. We conclude that these gaps might be of importance not only for molecular transport but also for cell passage across the vessel wall, which may be significant for tumor metastasis. VEGF may act as a paracrine effector to influence the permeability of lymphatic vessels via Flt-1, and of blood vessels via KDR.     

Olfactory control of behavior in rodents.

Examines the behavioral literature for information that could clarify the manner in which rodent behavior is mediated by olfaction. Functional use of olfaction is readily apparent in some of the behaviors reviewed. These behaviors include bait shyness; olfactorally-modulated aggression; olfactory marking of and recognition of stressful environments; and inhibition of food-seeking by odors generated from frustration. Other behaviors are reviewed in which olfaction is thought to play an important, though yet undelineated role, e.g., differences in approach preference based on sexual dimensions. Still other olfactory-associated behaviors are reviewed which meet current standards of inferential significance yet lack any apparent function for the animal. It is concluded that a coherent understanding of the functional use of olfaction by rodents awaits a shift in the philosophy of research away from rigorous examination of loosely selected odorants and behavioral measures toward rigorous examination of normal behaviors in natural social contexts. (80 ref.) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Photic entrainment of circadian rhythms in rodents

Photic entrainment of circadian rhythms occurs as a consequence of daily, light-induced adjustments in the phase and period of the suprachiasmatic nuclei (SCN) circadian clock. Photic information is acquired by a unique population of retinal photoreceptors, processed by a distinct subset of retinal ganglion cells, and conveyed to the SCN through the retinohypothalamic tract (RHT). RHT neurotransmission is mediated by the release of the excitatory amino acid glutamate and appears to require the activation of both NMDA- and non-NMDA-type glutamate receptors, the expression of immediate early genes (IEGs), and the synthesis and release of nitric oxide. In addition, serotonin appears to regulate the response of the SCN circadian clock to light through postsynaptic 5-HT1A or 5-ht7 receptors, as well as presynaptic 5-HT1B heteroreceptors on RHT terminals.     

An in vitro system for the study of matrix metalloproteases during decidualization in the mouse

Decidualization results in the remodeling of the extracellular matrix with the loss of collagen type I and the appearance of basement membrane matrix components. We have developed an in vitro assay system to study matrix metalloproteases during mouse decidualization. Uterine stroma, or decidua isolated from day 7.5 pregnant mice, were grown on a three-dimensional collagen type I matrix (Vitrogen). Gelatin zymography of conditioned media from these cultures showed constitutive secretion of processed forms of gelatinase A at 65, 62, and 59 kDa with 62 kDa predominating. Similar patterns of gelatinase A expression were obtained from tissue lysates of decidualizing uteri from days 5.5 to 7.5 of development. Cells cultured on Vitrogen, but not on plastic or matrix-coated dishes, were able to process the proenzyme to the 59 kDa form as observed in vivo. Only stroma cells cultured on a coating of collagen type I displayed the same increase in the 59 kDa zymogen. Decidua cells grown on Vitrogen attached and then migrated into aggregates that eventually penetrated the gel and spread as differentiated decidua on the underlying plastic. These preliminary results suggested that the in vitro assay system can be used to study the role of metalloproteases in matrix remodeling during decidualization.     

Proconvulsant and anticonvulsant effects of Evans blue dye in rodents

The effect of i.c.v. administration of Evans blue to sound sensitive DBA/2 mice and to genetically epilepsy-prone rats was studied. In mice, Evans blue (3.3-52 nmol) induced: hyperlocomotion, wild running, scratching, clonic muscle spasms, tonic seizure (latency 10-45 min), followed by death or recovery. The CD50 value for clonic seizures for Evans blue was 35(23-53) nmol. Pretreatment (45 min) with Evans blue (13-52 nmol, i.c.v.) dose-dependently reduced the incidence of sound-induced seizures in DBA/2 mice (ED50 value against clonic seizures = 30 [15-58] nmol, i.c.v). In rats, Evans blue (104 nmol, i.c.v.) induced electroencephalographic seizures in the hippocampus and cortex and behavioural limbic seizures with a latency of 15-20 min. A reduction in the mean score (from 5 to 2-3) for behavioural seizures was observed which lasted for 4-5 days in rats electrically-kindled daily in the hippocampal CA3 subsector. Sound-induced clonic seizures in kindled and non-kindled rats were reduced for 3-4 days after administration of Evans blue (104 nmol, i.c.v.).     

Matrix metalloproteinase and matrix metalloproteinase inhibitor expression in endometrial stromal cells during progestin-initiated decidualization and menstruation-related progestin withdrawal

Estradiol (E) primes human endometrial stromal cells (HESCs) for the decidualizing effects of progesterone in vivo and in vitro. Matrix metalloproteinase (MMP) expression was evaluated in confluent HESCs incubated in control medium, and in medium supplemented with either E, or the synthetic progestin medroxyprogesterone acetate (P), or E + P. Measurements with a specific ELISA indicated that basal pro-MMP-1 output was unaffected by E, whereas E + P, which induces the expression of several decidualization-related markers, produced a time-dependent inhibition in HESC-secreted levels of pro-MMP-1. Consistent with progestin inhibition of MMP-1 protein expression in the HESCs, P but not E, reduced steady state levels of MMP-1 messenger RNA (mRNA) as determined by Northern analysis. By contrast, mRNA levels for MMP-2 and the MMP inhibitor TIMP-1 were not altered by either P or E. Steroid withdrawal studies indicated that after MMP-1 expression was suppressed by incubation of the HESCs with E + P, 4 days of exposure to the antiprogestin RU 486 (mifepristone) significantly up-regulated MMP-1 levels in the conditioned medium by severalfold compared with cultures maintained in E + P. The change to steroid-free control medium required a more prolonged period of withdrawal to attain up regulatory effects that were comparable with those evoked by RU 486. The ELISA measurements were validated by immunoblot analysis with a specific MMP-1 antibody, which showed corresponding changes in a band at the expected mobility of about 50 kDa. Moreover, Northern analysis revealed parallel changes in MMP-1 mRNA levels, whereas neither MMP-2 nor TIMP-1 mRNA levels were modulated by adding or withdrawing steroids. The contrast between regulated MMP-1 expression and constitutive MMP-2 expression observed in the cultured HESCs is consistent with the demonstrated presence on the MMP-1 promoter of regulatory elements such as AP-1 and PEA-3 that are absent from the MMP-2 promoter. Extrapolation of these in vitro changes in HESCs to in vivo endometrial events suggests that: 1) inhibition of MMP-1 expression by E and progesterone would stabilize the perivascular endometrial ECM to prevent local hemorrhage during endovascular invasion by the implanting trophoblast; 2) enhanced expression of MMP-1 evoked by steroid withdrawal would mediate endometrial ECM degradation leading to sloughing of the functional layer during menstruation.     

A simple method for quantifying tremor in rodents

A simple and accurate device to evaluate frequency and intensity of involuntary tremor is described. Discrete and quantifiable measures of tremor could be obtained in terms of vertical (i.e., absolute effects at a given time) and horizontal (i.e. temporal) changes. The technique was evaluated employing dosages of physostigmine 0.1-0.7 mg/kg. Applications to other behavioral indices, such as locomotor activity and wet-dog shakes, are discussed.