Melatonin production by the guinea pig pineal gland in vitro

The fluid bathing pineal glands isolated from guinea pigs was collected serially and its melatonin content was estimated. Production was found to be high (500 pg/5 min) soon after isolation of the gland: it declined exponentially to a maintained lower level (50 pg/5 min) with a half-time of 16-20 min. A rapid increase could be produced by exposure of the gland to the beta-adrenoceptor agonist orciprenaline or by stimulating it electrically. The time course and the extent of the responses to either form of stimulation were similar in glands that had been taken in the morning, at dusk or in mid-dark: the prestimulation peak was lower from glands taken at dusk. Thus the pineal gland of the guinea pig is capable of responding rapidly to stimulation of its beta-adrenoceptors at any time. These responses parallel the depolarization observed intracellularly in this species when the pineal gland is stimulated electrically or exposed to beta-adrenoceptor agonists.     

Relaxin has a minor role in rat mammary gland growth and differentiation during pregnancy

Growth and differentiation of mammary gland is associated with numerous hormones and a variety of cell-cell, cell-matrix interactions. This study addressed the role of relaxin (Rlx) on these processes. Morphologic and biochemical changes that occur throughout the second half of pregnancy are reported. Temporal patterns and spatial distributions of markers useful to evaluate proliferation, secretion, and collagen remodeling were established. To evaluate the role of Rlx, an ablation/replacement animal model was used. Considering Rlx secretion pattern, two periods were selected: d 11 through d 13, and d 20 through d 23. In the stroma, the extracellular compartment showed changes associated with the lack of Rlx. Collagen remodeling within the lobuloalveolar structure, measured by a significant increase in collagen birefringence, decreased at d 12, d 21, and d 22. Parenchymal structures were less sensitive to the absence of Rlx than stroma. Epithelial cell proliferation was lower in Rlx-deficient rats only at d 12, and α-lactalbumin expression decreased at d 21 and d 22. Both lobuloalveolar diameter and percentage of area occupied by these structures showed no changes. In the absence of Rlx, some of the studied markers showed statistically significant differences in scattered days; these do not make clear trends. No differences were found on d 23 on any of the studied parameters suggesting that compensatory mechanisms might be activated to overcome the effects of the absence of Rlx. Unlike the critical role of Rlx either in uterine cervix dilation or in nipple development during rat pregnancy, Rlx had a minor role in growth and differentiation of rat mammary gland.     

Substance P immunoreactivity in the anterior pituitary gland of the guinea pig

The aim of this study was to demonstrate the presence of Substance P (SP) immunoreactive cells within the anterior pituitary gland of the adult guinea pig. By utilizing the PAP technique of Sternberger, immunoreactive SP-containing cells were visualized in all animals studied. These cells were most dense in the ventral portion of the gland. When adjacent tissue sections were incubated with anti-SP serum and anti-rat TSH serum, it was observed that most, if not all, the SP immunoreactive cells examined also exhibited TSH immunoreactivity. However, not all TSH immunoreactive cells contained SP immunoreactivity. Immunoabsorption controls indicated that the immunohistochemical staining reactions were specific for both SP and TSH. The results of this study indicate that immunoreactive SP is present in the guinea pig anterior pituitary and that this peptide is localized within TSH immunoreactive cells.     

Relaxin production and release after hysterectomy in the pig

Relaxin secretion by luteal tissue and into peripheral blood was examined in relation to changes in the fine structure of aging porcine corpora lutea. Sequential bleedings every sixth day reveal a similar increase in serum relaxin concentrations from days 6-114 in hysterectomized and pregnant gifts. The span of the estrous cycle is about 21 days, and the length of pregnancy is about 115 days in pigs. Estrone (E1) and 17 beta-estradiol (17 beta-E2) serum levels increase with placental development; after hysterectomy, they are low from days 12-168. In luteal tissue, peak quantities of relaxin and electron-dense granules occur on day 100 in hysterectomized and pregnant gilts. By day 112, luteal tissue relaxin levels and granule populations decrease by about half, while relaxin secretion into peripheral blood approaches peak concentrations about day 114 in hysterectomized as well as pregnant gilts. Between days 114 and 120, serum relaxin decreases by half even though the corpora lutea persist to day 150 in hysterectomized animals. Thereafter, cytoplasmic granules and blood and luteal tissue levels of relaxin decrease gradually to day 150 in hysterectomized gilts; in contrast, they disappear abruptly after parturition. In hysterectomized gilts, relaxin release coincides with diminished populations of granules in corpora lutea, and the initiation of these events seems to be precisely timed. E1 and 17 beta-E2 secretion is unrelated to relaxin secretion in pregnant and hysterectomized gilts. High peripheral blood levels of E1 and 17 beta-E2, primarily of placental origin, or even the presence of the uterus are not required in the release of relaxin at day 114. By using steroidogenic and protein synthetic capabilities of luteal cells as criteria essential to the evolution of reproduction in this species, the results presented here suggest that 114 days rather than 21 days characterize the reproductive cycle, even in the absence of the uterus. These results indicate that the production and, particularly, the release of relaxin into peripheral blood on about day 114 in hysterectomized gilts as well as pregnant animals may be genetically controlled.     

Two-dimensional analysis of ventricular fibrillation in the guinea pig

Cardiac arrhythmias are undesirable electrical activity in the heart. Ventricular fibrillation (VF) is a fatal cardiac arrhythmia and is characterized by the breakdown of organized electrical activity in the ventricular myocardium. However, little is known about VF, partially because it is difficult to study and understand an apparently disorganized activity. One method for discovering the nature of VF is processing the epicardial electrical signals by using cardiac mapping techniques. These techniques involve the study of propagation patterns seen in VF. In this project, we used guinea pigs to study the spatial organization of the epicardial electrical activity during VF. VF was induced in 9 open chest guinea pigs and epicardial electrode data were acquired by using a square array of 192 electrodes for 4 seconds every minute. The mean correlation length and dominant frequency were measured in each recorded segment. Correlation length is a measure of the spatial order in a system and dominant frequency is the frequency corresponding to the spectral maximum. The mean correlation length was found to vary between 1-7 mm and the dominant frequency was in the range of 1-14 Hz. This study suggests that guinea pig VF exhibits a level of organization equivalent to or greater than that seen in previous studies.     

Identification of mouse mammary epithelial cells by immunofluorescence with rabbit and guinea pig antikeratin antisera.

Few markers are available to identify the three types of mammary epithelial cells--ductal, alveolar, and myoepithelial--especially in pathological conditions and in cell cultures. We have used antisera to human keratins in immunofluorescence to facilitate the identification of the three mouse mammary epithelial cell types. In frozen tissue sections and primary cell cultures, a rabbit antikeratin antiserum specifically stained cytoplasmic filaments in all three types of epithelial cells. A guinea pig antiserum against the same keratin preparation, however, reacted preferentially with filaments in myoepithelial cells and readily detected this cell type in normal, dysplastic, and malignant mammary tissues and cell cultures. Neither antisera reacted with fibroblasts or any other mesenchymal cells. The combined use of the two antikeratin antisera thereby permits rapid surveys of tissue sections and cultures for the localization of not only all epithelial cells but also the subpopulation of myoepithelial cells. Moreover, when mammary cultures established from late-pregnant or lactating mice were stained simultaneously with guinea pig antikeratin and rabbit anticasein antisera, three populations of epithelial cells were mutually exclusive: those stained by anticasein antiserum, those stained by guinea pig antikeratin antiserum, and those stained by neither, consistent with properties of alveolar, myoepithelial, and ductal cells, respectively. These antisera thus offer a tool for studying different epithelial cell types during mammary development, tumorigenesis, and malignant progression.     

The association between melatonin production and electrophy siology of the guinea pig pineal gland

Abstract: Melatonin production by isolated pineal glands from guinea pigs was examined under conditions that affect membrane potential or the firing of action potential-like spikes. In glands from superior cervical ganglionectomized animals, depolarization resulting from increasing extracellular potassium concentration to 100 mM did not initiate melatonin production, and it delayed the response to the β-adrenoceptor agonist orciprenaline. In glands from intact animals, melatonin production was initiated by exposure to 100 mM potassium with a time-course similar to the response to orciprenaline. A proportion of this response was propanol resistant, suggesting that the normal control of melatonin production may involve a neurotransmitter in addition to norepinephrine. Exposure to verapamil or nifedipine, or removal of extracellular calcium, previously shown to eliminate action potential-like spikes, did not substantially affect the increase in melatonin production induced by orciprenaline. Phenylephrine, which stimulates spiking, produced only a slight increase in melatonin production. It is concluded that the depolarization and the spiking are not closely related to the stimulation of melatonin production, but may relate principally to the secretion of a substance other than melatonin.     

Relaxin and progesterone secretion as affected by luteinizing hormone and prolactin after hysterectomy in the pig

Plasma levels of relaxin and progesterone in hysterectomized and pregnant gilts were determined from days 100-120 to evaluate the effects of purified porcine (p) LH and pPRL on the secretory activity of the aging corpora lutea. Gilts were bred on the second observed estrus or were hysterectomized between 6-8 days after estrus (estrus = day 0) and were assigned randomly to one of three treatment groups; saline-treated control, im injections of pLH, and iv injections of pPRL from days 110-120. In control, pLH-treated, and pPRL-treated animals, average gestation lengths were 114 +/- 0.8, 116 +/- 1.9, and 115 +/- 0.5 days (+/- SE), respectively. The relaxin level in mated gilts on day 100 was less than 2 ng/ml; it began to increase after day 110 and peaked in control animals on day 113 (66 ng/ml), whereas in pLH- and pPRL-treated animals, prepartum peak values were greater (P less than 0.01) and occurred on days 113 (104 ng/ml) and 114 (117 ng/ml), respectively. Relaxin dropped to basal levels (less than 1 ng/ml) by day 115 in controls and by day 116 in both pLH- and pPRL-treated gilts. Although pLH and pPRL treatments markedly accentuated peak relaxin secretion, they did not significantly accelerate or delay parturition or delay the abrupt demise of the corpora lutea immediately postpartum. In hysterectomized gilts, relaxin began to increase after day 110, peaked in control animals on day 113 (27 ng/ml), and decreased abruptly thereafter to less than 4 ng/ml. In contrast, pLH caused an immediate release of relaxin on day 111 (23 ng/ml) and sustained elevated levels (P less than 0.01) of relaxin until day 118, but the original corpora lutea regressed. Relaxin in pPRL-treated animals increased steadily after day 110, reaching peak values by day 115 (29 ng/ml), and remained consistently elevated (P less than 0.01) until day 120. Progesterone secretion was maintained in the pPRL-treated hysterectomized gilts from days 110-120 by the original corpora lutea and with no luteinization of follicles or formation of new corpora lutea. It is evident from this study that administration of pPRL starting on day 110 enhanced and prolonged the preprogramed release of relaxin and maintained progesterone secretion by aging corpora lutea in hysterectomized animals until day 120.     

Forskolin stimulation of naphthylamidase in guinea pig thyroid sections detected with a cytochemical bioassay

Forskolin, from the roots of the Indian medicinal plant Coleus forskohlii, has recently been shown to be a potent stimulator of adenylate cyclase in many systems, including endocrine tissues such as the thyroid gland. We describe forskolin activation of beta-naphthylamidase activity in guinea pig thyroid tissue using the cytochemical bioassay (CBA) for thyroid stimulators. This CBA is the most sensitive bioassay for TSH and LATS-B currently available, being able to detect stimulation by doses as low as 10(-5) mU TSH/l and 10(-9) mU LATS-B/l. The dose-response curve to forskolin was bell-shaped (as is seen with TSH and LATS-B) with the ascending limb of the curve produced by 10(-13) M to 10(-12) M forskolin after a 3 min exposure time. Maximal stimulation was observed with 10(-12) M forskolin. However, the dose-response curve to forskolin was not parallel to that given by TSH, the slope of the ascending limb being much greater. It has been suggested that stimulation of beta-naphthylamidase activity in the CBA is via cAMP. We report that dibutyryl cAMP at doses from 10(-16) M to 10(-11) M produces a bell-shaped dose-response curve with a very broad peak response, again not parallel to that produced by TSH. Forskolin activation of beta-naphthylamidase in the CBA is unaffected by a 1:10(6) dilution of 11E8, a monoclonal antibody raised against solubilised TSH receptors, which binds to the TSH receptor and inhibits TSH stimulation. Although the precise location of forskolin action is not known, this is further evidence that forskolin acts at a post-surface receptor site.     

哮喘豚鼠气道重塑与气道反应性的图像分析

目的研究哮喘豚鼠气道重塑的机制及气道反应性的变化。方法实验分二组对照组（20只）和哮喘组（20只）,取双肺与不同刺激因素作用后作组织切片、HE染色,通过图像分析仪测定支气管内周长、管壁厚度、外周长,并计算平滑肌收缩百分比（PMS）。结果（1）哮喘组支气管壁厚度（WA/Pi）为（10.0±2.0）μm     

Suppression of lymphoproliferation by alveolar macrophages in the guinea pig

Setting : The relationship between alveolar macrophages (AM) and lymphocytes may be important in the early establishment of infection with Mycobacterium tuberculosis . AM in several species have been shown to suppress lymphoproliferation by producing inhibitors that include nitric oxide (NO).Objective : To study this phenomenon in the guinea pig, the mitogen-induced proliferation of splenic lymphocytes was quantified under various conditions of co-culture with resident AM.Results : Guinea pig AM consistently and profoundly suppressed proliferation in the co-cultures at AM:lymphocyte ratios of 1:4 or greater. The inclusion of a NO synthesis inhibitor, N-monomethyl-L arginine (NMMA), in the co-culture medium did not influence the suppression of Con A-induced lymphoproliferation by resident guinea pig AM. No nitrite could be detected in supernatant fluids of co-cultured AM and splenocytes. Attempts to stimulate guinea pig AM with LPS in combination with recombinant murine and human IFN--γ, infection with live Listeria monocytogenes , or incubation with the supernatants from ConA-activated guinea pig lymphocytes failed to generate NO metabolites. The addition of catalase or indomethacin to the Con A-induced AM-splenocyte co-cultures, to inhibit hydrogen peroxide (H₂O₂) or prostaglandin E₂(PGE₂), respectively, did not counteract the suppression mediated by AM. Cell contact was necessary for the co-cultures to generate their inhibitory effects on lymphoproliferation, however, the suppression was actually mediated, at least in part, by soluble factors produced in the co-cultures.Conclusion : These results suggest that resident alveolar macrophages suppress lymphocyte proliferation in the guinea pig, but that the effect is not mediated by NO, PGE₂or H₂O₂. The failure to demonstrate NO synthesis under a variety of stimulatory conditions, which resulted in macrophage activation, suggests that the guinea pig is similar to the human in that regard.     

Collagenase production by guinea pig megakaryocytes in vitro

Cultured guinea pig bone marrow megakaryocytes were found to secrete a 92-kd collagenase that was detected by digestion of gelatin in a polyacrylamide substrate gel assay. Neither casein or bovine serum albumin were digested by this enzyme. The enzyme is a neutral metalloprotease. Its secretion is increased by thrombin (1.0 U/ml) and phorbol myristate acetate (10 ng/ml) and is unaffected by prostaglandin E1 (10 microM). In the absence of serum, gelatinase secretion is inhibited, but it can be stimulated by cytochalasin D (1.0 microgram/ml). Gelatinase activity in the medium from megakaryocytes cultured on rat tail collagen gel is decreased. Medium from megakaryocytes cultured on Matrigel contains a second gelatinase of 90 kd. Addition of the tetrapeptide RGDS to the cultures on Matrigel blocks the appearance of the 90-kd gelatinase. Platelets contained both a 92- and a 90-kd gelatinase that was detected only after thrombin activation. The results indicate that megakaryocytes can secrete a collagenase, and its secretion may be in part controlled by interaction with the extracellular matrix. The appearance of the 90-kd gelatinase may be associated with megakaryocyte maturation and platelet formation.     

Propagation by sporulation in the guinea pig symbiont Metabacterium polyspora

The Gram-positive bacterium Metabacterium polyspora is an uncultivated symbiont of the guinea pig gastrointestinal tract. Here we present evidence that in M. polyspora vegetative cell division has taken on a minor, and apparently dispensable, role in propagation. Instead, this unusual bacterium has evolved the capacity to produce progeny in the form of multiple endospores. Endospore formation is coordinated with transit of the bacterium through the gastrointestinal tract of the guinea pig. For the majority of cells, sporulation is initiated in the ileum, whereas later stages of development take place in the cecum. We show that multiple endospores are generated both by asymmetric division at both poles of the cell and by symmetric division of the endospores at an early stage of their development. Our findings suggest that M. polyspora represents an intermediate step in the evolution of a novel mode of cellular propagation that originates with endospore-forming Bacillus and Clostridium spp., which reproduce by binary fission, and extends to Epulopiscium spp., which create multiple viviparous offspring by a process of internal reproduction.     

Development of the mammary gland and lactation

Many hormones, growth factors, and protooncogene products involved in mammary development and lactation have been identified; however, the mechanism of their concerted action remains to be explained. In addition to these regulatory factors, normal mammary development and lactation require the cell-cell interaction of stromal and parenchymal elements in the mammary gland. Recent studies indicate that PRL effects on target cells are likely transmitted into cells via activation of tyrosine kinase associated with a protein-designated JAK-2, while other studies have identified stimulatory and inhibitory factors that interact with the 5' promotor regions of milk-product genes.