Biomedical subjects

K F Hauser

Publications and source records attributed to K F Hauser.

At least 55 records · Page 3Linked to original sources

PubMed1993

Endogenous opioid systems and the growth of oligodendrocyte progenitors: paradoxical increases in oligodendrogenesis as an indirect mechanism of opioid action.

Endogenous opioids inhibit nervous system development by inhibiting the proliferation of certain neuronal and glial progenitors. To determine whether opioids affect the growth of preoligodendrocytes, the effects of the endogenous opioid [Met5]-enkephalin were examined in preoligodendrocytes in primary mixed-glial and preoligodendrocyte-enriched (> 98% pure) cultures. Proliferating preoligodendrocytes in mixed-glial or preoligodendrocyte-enriched cultures were continuously treated for a total of 40 h with either basal growth media (controls), 1 microM [Met5]-enkephalin, 1 microM [Met5]-enkephalin plus the opioid antagonist naloxone (3 microM), or naloxone alone (3 microM), and incubated in [3H]-thymidine (0.2 microCi/ml/4-6 h) after 34-36 h of opioid exposure. Opioid-dependent changes in DNA synthesis were assessed autoradiographically in O4-immunoreactive oligodendrocyte progenitors. Naloxone alone significantly decreased the rate of DNA synthesis and number of O4-immunoreactive preoligodendrocytes in mixed-glial cultures. However, naloxone and/or [Met5]-enkephalin did not affect DNA synthesis or the number of O4-immunoreactive preoligodendrocytes in cultures enriched in preoligodendrocytes. The results suggest that astrocytes, or perhaps another cell type, play a permissive role in opioid-dependent alterations in preoligodendrocyte proliferation. Endogenous opioids affect the genesis of neural cells by both direct and indirect mechanisms.

Animals↗

PubMed1993

Survival of extraocular muscle in long-term organotypic culture: differential influence of appropriate and inappropriate motoneurons.

The myotrophic effects of a precise matching of motoneuron to target muscle may be particularly apparent during the development of some unique skeletal muscle types, such as the extraocular muscles. To understand how motoneuron-specific factors modulate muscle ontogeny, neonatal extraocular muscle explants were cocultured with either the appropriate midbrain motoneurons or the inappropriate spinal cord motoneurons. Thigh muscle cocultured with spinal motoneurons, which differentiates and survives for several months, served as a control. In short-term cultures (during the first 3 weeks in vitro), neonatal extraocular muscle explants developed myotubes that were immunoreactive for myosin, became innervated, and matured in parallel with hindlimb muscle explants. The origin of the motoneurons (midbrain or spinal cord) did not affect extraocular muscle development during the first 3 weeks in vitro. However, in long-term cultures (after the third week in vitro), extraocular muscle that was innervated by the inappropriate spinal motoneurons failed to survive and degenerated. The failure of extraocular muscle to survive when cocultured with the inappropriate motoneurons did not result from an artifact of in vitro conditions. When extraocular muscle primordia were innervated by midbrain explants, many of which contain oculomotor motoneurons, the cultures thrived for > 60 days. We conclude that the trophic requirements of extraocular muscle in organotypic culture are different from those of skeletal muscle isolated from the hindlimb. The novel oculomotor motoneuron-specific interactions that are essential for the survival of extraocular muscle primordia in vitro may prove to be important for the determination of the unusual properties of extraocular muscle in vivo.

Animals↗

PubMed1993

Diversity and developmental regulation of extraocular muscle: progress and prospects.

The developmental regulation of skeletal muscle phenotypes has attracted much attention from cell and molecular biologists. Myogenesis serves as an excellent model of the interaction of genetic and epigenetic events in determining definitive adult characteristics. The muscles that are responsible for eye movements are among the most structurally and functionally diverse mammalian skeletal muscles. Despite these unique attributes, the extraocular muscle has been studied less extensively than the traditional limb and diaphragm models. This review explores current concepts regarding the development of these muscles. Proper interocular alignment and coordination of eye movements are essential for normal vision in the adult. Moreover, coordinated oculomotor function during the perinatal period is critical for the normal development of the visual system. Therefore, a thorough understanding of the sequence of events and mechanisms that regulate extraocular muscle development and maldevelopment is vital. Studies related to myoblast origin and differentiation and to the role of genetic versus environmental factors in shaping and maintaining adult extraocular muscle phenotype are reviewed. Prospects for understanding the cellular and molecular mechanisms that regulate the heterogeneity and plasticity of these muscles also are discussed.

Animals↗

PubMed1992

Morphine regulates DNA synthesis in rat cerebellar neuroblasts in vitro.

The effects of morphine on DNA synthesis by external granular layer (EGL) neuroblasts was examined in whole-mount organotypic cultures isolated from 10-day-old rat cerebella using bromodeoxyuridine (BrdU). After 24 h in vitro, explants were treated for 24 h with 10 nM, 1 or 100 microM morphine, morphine plus 30 nM, 3 or 300 microM of the opiate antagonist naloxone, respectively, or those concentrations of naloxone alone. BrdU was added during the last 4 h of drug treatment. EGL neuroblasts were unambiguously identified by size and morphology, location and by protein kinase C II immunocytochemistry. The proportion of EGL neuroblasts incorporating BrdU was significantly reduced in the presence of 1 microM morphine, while 100 microM morphine had little additional effect. The concentration of morphine predicted to cause a half-maximal reduction in BrdU labeling index was 22.5 nM. Morphine's ability to reduce BrdU incorporation by EGL neuroblasts was concentration dependent and was prevented by concomitant treatment with naloxone, implicating the involvement of opioid receptors. The results suggest that morphine can directly regulate the growth of the developing cerebellum by inhibiting neuroblast proliferation within the EGL.

Animals↗

PubMed1992

Cell proliferation and protooncogene induction in oligodendroglial progenitors.

Cell proliferation and the expression of the protooncogenes c-fos and c-jun have been examined in the primary cultures of oligodendroglial (OL) progenitor cells in response to phorbol 12-myristate 13-acetate (PMA), serum, insulin, insulin-like growth factor-I (IGF-I), platelet-derived growth factor (PDGF), and fibroblast growth factor (FGF). Combined [3H]thymidine autoradiography and immunocytochemistry was used to assess the mitogenic response of O4 (an oligodendrocyte-specific marker)-positive OL progenitors. In addition, the rate of DNA synthesis was measured by the incorporation of [3H]thymidine into acid-precipitable material. It was found that all of the agents tested stimulated DNA synthesis in OL progenitors and induced a rapid increase in c-fos and c-jun protooncogene expression. The induction of c-fos gene expression and DNA synthesis in response to PMA was completely blocked by 1-(5-isoquinolinyl-sulfonyl)-2-methylpiperazine (H-7), a potent inhibitor of protein kinase C (PKC), thereby suggesting a role for PKC in the control of c-fos expression and cell proliferation in OL progenitors.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1991

Expression of proenkephalin mRNA in developing cerebellar cortex of the rat: expression levels coincide with maturational gradients in Purkinje cells.

The cellular localization of proenkephalin (PE) mRNA expression was systematically examined in midsagittal (vermal) sections of the developing rat cerebellar cortex by in situ hybridization. PE mRNA was initially detected in Golgi cells of postnatal day 7 (PND 7) rats and in each group thereafter. Moreover, PND 7 rats also displayed an intense layer of PE mRNA hybridization signal over the Purkinje cell layer. By PND 14, distinct cellular labeling was observed in a subpopulation of Purkinje cells in all lobules of the vermis except lobule III. At PND 7 and 14, the area and level of intensity of Purkinje cell associated PE mRNA hybridization signal followed a gradient that was most intense caudally but then decreased rostrally. At PND 21, the proportion of labeled Purkinje cells and the intensity of PE hybridization signal was evenly dispersed between the anterior and posterior lobules of the cerebellar vermis. PE hybridization signal was not detected in the developing neural cells of the external granular layer or the interneurons of the molecular layer in the vermis. These results indicate that the ontogeny of PE mRNA expression in Purkinje cells is developmentally regulated since levels of expression closely follow the chronological order of settling and maturation of these neurons. Based on prior evidence that endogenous opioids inhibit the growth of Purkinje cell dendrites and dendritic spines, PE expression is likely to be important for Purkinje cell maturation.

Animals↗

PubMed1991

Characterization of opioid-dependent glial development in dissociated and organotypic cultures of mouse central nervous system: critical periods and target specificity.

Opioid-dependent changes in glial growth were characterized in primary dissociated and organotypic explant cultures of the developing mouse central nervous system (CNS) continuously grown in the presence of an endogenous opioid, [Met5]enkephalin, or an opiate drug, morphine. The glia in dissociated, astrocyte-enriched cultures derived from the cerebra of postnatal day 1, 3, or 5 mice, respectively, displayed age-dependent reductions in glial numbers that occurred at 3, 7, or 9 days in vitro (DIV) in response to continuous [Met5]enkephalin (10(-6) M) exposure. In contrast, in cultures derived from gestational day 19 mice, glial numbers were not reduced following continuous exposure to 10(-6) M [Met5]enkephalin during the first 7 days in vitro. An examination of [3H]thymidine incorporation by glial fibrillary acidic protein-(GFAP) immunoreactive astrocytes with flat (type 1) morphology in dissociated cultures derived from postnatal day 1 mice revealed that the reduction in glial numbers at 3 DIV was not immediately preceded by a reduction in the rate of [3H]thymidine incorporation at 2 DIV, although previous studies have shown that opioids inhibit the rate of [3H]thymidine incorporation by more mature astrocytes at 4 or 6 DIV. Early (i.e., at 2 to 3 DIV) changes in glial numbers may result from an inhibition of the proliferative rate of non-GFAP-containing glia or astrocyte precursors, or an enhanced rate of glial death. The rate of [3H]thymidine incorporation by GFAP-immunoreactive astrocytes with process-bearing (type 2) morphology was unchanged by opioid treatment. In separate experiments, a comparison of the area of growth of GFAP-immunoreactive astrocytes in paired symmetrical (right vs left) organotypic explant cultures demonstrated that opiates (i.e., 10(-5) M morphine) can inhibit astrocyte growth when the normal histiotypic organization of neurons and glia are maintained, and that there are regional differences in astrocyte responsiveness. Opioid-dependent alterations in astrocyte growth were mediated through specific opioid receptors since they were prevented by simultaneous treatment with (-)naloxone. The results suggest that the ability of opioids to modify glial growth is highly selective and varies depending on astrocyte type, as well as temporal and regional factors. Spatial and temporal differences in the response of developing glia to opioids may determine critical periods of CNS vulnerability to opioids in the maturing brain.

Animals↗

PubMed1991

Morphine alters astrocyte growth in primary cultures of mouse glial cells: evidence for a direct effect of opiates on neural maturation.

To determine whether exogenous opiate drugs with abuse liability directly modify neural growth, the present study investigated the effects of morphine on astrocyte proliferation and differentiation in primary cultures of murine glial cells. The results indicate that morphine decreases glial cell production in a dose-dependent, naloxone-reversible manner. Most notably, gliogenesis virtually ceased in the presence of 10(-6) M morphine during the first week in culture, whereas 10(-8) M or 10(-10) M morphine caused an intermediate suppression of growth compared to control or 10(-6) M morphine treated cultures. Moreover, morphine treatment inhibited [3H]thymidine incorporation by glial fibrillary acidic protein (GFAP) immunoreactive, flat (type 1) astrocytes, suggesting that the decrease in glial cell production was due in part to an inhibition of astrocyte proliferation. Morphine also caused significant increases in both cytoplasmic area and process elaboration in flat (type 1) astrocytes indicating greater morphologic differentiation. In the above experiments, morphine-dependent alterations in astrocyte growth were antagonized by naloxone, indicating that morphine action was mediated by specific opioid receptors. These observations suggest that opiate drugs can directly modify neural growth by influencing two critical developmental events in astrocytes, i.e., inhibiting proliferation and inducing morphologic differentiation.

Animals↗

PubMed1991

Glial growth is regulated by agonists selective for multiple opioid receptor types in vitro.

To determine whether one or more opioid receptor types might be preferentially involved in gliogenesis, primary mixed glial cultures derived from mouse cerebra were continuously treated with varying concentrations of opioid agonists selective for mu (mu), i.e., DAGO ([D-Ala2, MePhe4, Gly(ol)5]enkephalin), delta (delta), i.e., DPDPE ([D-PEN2,D-PEN5]enkephalin), or kappa (kappa), i.e., U69,593, opioid receptor types. In addition, a group of cultures was treated with [Met5]-enkephalin, an agonist for delta opioid receptors as well as putative zeta (zeta) opioid receptors. Opioid-dependent changes in growth were assessed by examining alterations in (1) the number of cells in mixed glial cultures at 3, 6, and 8 days in vitro (DIV), (2) [3H]thymidine incorporation by glial fibrillary acidic protein (GFAP) immunoreactive, flat (type 1) astrocytes at 6 DIV, and (3) the area and form factor of GFAP-immunoreactive, flat (type 1) astrocytes. DPDPE at 10(-8) or 10(-10) M, as well as [Met5]-enkephalin at 10(-6), 10(-8), or 10(-10) M, significantly reduced the total number of glial cells in culture; but this effect was not observed with DAGO or U69,593 (both at 10(-6), 10(-8), or 10(-10) M). Equimolar concentrations (i.e., 10(-6) M) of [Met5]enkephalin or U69,593, but not DPDPE or DAGO, suppressed the rate of [3H]thymidine incorporation by GFAP-immunoreactive, flat (type 1) astrocytes. DAGO had no effect on growth, although in previous studies morphine was found to inhibit glial numbers and astrocyte DNA synthesis. [Met5]enkephalin (10(-6) M) was the only agonist to significantly influence astrocyte area. Collectively, these results indicate that delta (and perhaps mu) opioid receptor agonists reduce the total number of cells in mixed glial cultures; while [Met5]enkephalin-responsive (and perhaps kappa-responsive) opioid receptors mediate DNA synthesis in astrocytes. This implies that delta opioid receptors, as well as [Met5]enkephalin-sensitive, non-delta opioid receptors, mediate opioid-dependent regulation of astrocyte and astrocyte progenitor growth. These data support the concept that opioid-dependent changes in central nervous system growth are the result of endogenous opioid peptides acting through multiple opioid receptor types.

Analgesics↗

PubMed1991

Co-localization of proenkephalin mRNA using cRNA probes and a cell-type-specific immunocytochemical marker for intact astrocytes in vitro.

To determine whether cultured astrocytes express opioid gene mRNA, a method was developed for co-localizing a cell-type specific immunocytochemical marker for astrocytes, glial fibrillary acidic protein (GFAP), and proenkephalin mRNA in situ hybridization signal using high affinity cRNA probes. GFAP immunoreactivity and proenkephalin mRNA hybridization reaction were examined in intact glial cell preparations from neonatal mice that were cultured for 4-6 days prior to fixation. The double labelling method described herein permits the unambiguous identification of mRNA expression in specific populations of intact cultured cells using cell type-specific markers.

Animals↗

PubMed1990

Cellular localization of proenkephalin mRNA and enkephalin peptide products in cultured astrocytes.

To identify the possible cellular sites of opioid gene expression during ontogeny, proenkephalin mRNA and enkephalin peptide expression were examined, respectively, by in situ hybridization and immunocytochemistry in organotypic explants of rat cerebellum and in astrocyte-enriched cultures of murine cerebral hemispheres. High levels of proenkephalin mRNA and enkephalin immunoreactivity were detected in immature cells identified as astrocytes. Double-labeling studies combining in situ hybridization and immunocytochemical localization of the astrocytic marker, glial fibrillary acidic protein, provided direct evidence that proenkephalin mRNA is expressed by astrocytes in culture. Based on previous studies that Met-enkephalin can inhibit astrocyte growth in vitro, the present results suggest that proenkephalin gene expression by astrocytes is important during central nervous system maturation.

Animals↗

PubMed1990

Opioid-dependent growth of glial cultures: suppression of astrocyte DNA synthesis by met-enkephalin.

The action of met-enkephalin on the growth of astrocytes in mixed-glial cultures was examined. Primary, mixed-glial cultures were isolated from 1 day-old mouse cerebral hemispheres and continuously treated with either basal growth media (controls), 1 microM met-enkephalin, 1 microM met-enkephalin plus the opioid antagonist naloxone (3 microM), or naloxone alone (3 microM). Absolute numbers of neural cells were counted in unstained preparations, while combined [3H]-thymidine autoradiography and glial fibrillary acid protein (GFAP) immunocytochemistry was performed to identify specific changes in astrocytes. When compared to control and naloxone treated cultures, met-enkephalin caused a significant decrease in both total cell numbers, and in [3H]-thymidine incorporation by GFAP-positive cells with flat morphology. These results indicate that met-enkephalin suppresses astrocyte growth in culture.

Animals↗

PubMed1989

Androgen increases the number of cells in fetal mouse spinal cord cultures: implications for motoneuron survival.

Androgen effects were studied in organotypic cultures of the E12 fetal mouse lumbosacral spinal cord labeled in utero with [3H]thymidine on E10. Following continuous exposure to androgens for one month in vitro, the number of labeled cells was significantly increased in whole explants, and in hemisected segments in the presence or absence of co-cultured fetal thigh muscle. Because lumbosacral motoneurons undergo their final mitosis predominantly on E10 and thus remain permanently labeled, the results suggest that androgens increase neuronal numbers by directly modulating motoneuron survival rather than stimulating mitosis. These findings demonstrate for the first time that in addition to the well documented role of the muscle target in motoneuron survival, the direct neuronotrophic effects of androgen at the level of the spinal cord must also be considered.

Animals↗

PubMed1989

Endogenous opioid systems and the regulation of dendritic growth and spine formation.

The role of endogenous opioid systems (endogenous opioids and opioid receptors) in neuronal development was examined in 10- and 21-day-old rats by utilizing an opioid antagonist (naltrexone) paradigm. Throughout the first 3 weeks of life, Sprague-Dawley rats were given daily subcutaneous injections of either 50 mg/kg naltrexone, a dosage that invoked a complete (24 hours/day) receptor blockade, or 1 mg/kg naltrexone, a dosage which intermittently blocked (4-6 hours/day) opioid receptors and exacerbated opioid action; animals injected with sterile water served as controls. Pyramidal cells from the frontoparietal cortex (layer III) and hippocampal field CA1, and cerebellar Purkinje cells, were impregnated by using the Golgi-Kopsch method; total and mean dendrite segment length, branch frequency, and spine concentration were analyzed morphometrically. Perturbations of endogenous opioid systems caused region-dependent alterations in dendrite complexity and/or spine concentration in all brain areas. Continuous opioid receptor blockade resulted in dramatic increases in dendrite and/or spine elaboration compared to controls at 10 days in all brain regions; however, these increases were only evident in the hippocampus at 21 days. With intermittent blockade, dendrite and/or spine growth were often subnormal, being predominant at day 21. Our results indicate that endogenous opioid systems are critical regulators of neuronal differentiation, and they control growth through an inhibitory mechanism. Considering previous findings demonstrating that neurobehavioral ontogeny is dependent on endogenous opioid-opioid receptor interactions, the present results suggest an opioid-dependent, structure-function relationship between neuronal and behavioral maturation.

Animals↗

PubMed1988

Autoradiographic studies of cerebellar histogenesis in the premetamorphic bullfrog tadpole: II. Formation of the interauricular granular band.

This study examines the origin of cells in the interauricular granular band (iagb) in the cerebellum of the frog tadpole during early stages of development by means of histological and autoradiographic methods. Premetamorphic bullfrog tadpoles were exposed to multiple doses of 3H-thymidine (10 microCi/g body weight per exposure) at developmental stages ranging from 1 week to 1 year and were killed at either 6 or 12 months of age. The autoradiographic data were examined to determine the time when cells of the iagb were generated. Our findings show that initial generation of iagb cells begins at week 3 and that a peak in the formation of postmitotic neurons is reached at the age of 10 weeks. This is followed by other peaks of cell generation at the ages of 16 weeks, 10 months, and 11.5 months. The generation cycles of iagb cells are interrupted by periods of quiescence when label cannot be detected in any of the cells. These quiescent periods occur at the ages of 20-26 weeks, 7 months, and 12 months. These findings indicate that cells of the iagb are generated in a cyclical manner over the entire 1-year period which was studied. Comparison of our present data on iagb cell formation with the generation of cells in the EGL shows that the production of these two groups of cells is overlapping, but cells of the iagb begin and cease production before those of the EGL. On the basis of our findings we propose that the cells of the iagb and the EGL belong in separate cell groups which are generated by distinct subpopulations of germinal cells in the neuroepithelial cap.

Animals↗

PubMed1987

Autoradiographic studies of cerebellar histogenesis in the premetamorphic bullfrog tadpole: I. Generation of the external granular layer.

This study examines the time of origin of cells in the external granular layer (EGL) in the frog cerebellum during early stages of development. Premetamorphic bullfrog tadpoles were given multiple intraperitoneal injections of 3H-thymidine (10 microCi/g body weight per injection) at developmental stages ranging from 4 weeks to 1 year and were killed at either 6 or 12 months of age. Autoradiograms were analyzed to determine the time when cells of the EGL were generated by an examination of the labeling pattern in the neuroepithelial cap where EGL cells were presumably formed and in the EGL into which they migrated. The developmental stage of the cerebellum in the 6-month-old tadpole was essentially the same as that of the 12-month-old animal except for an increased size in the older tadpole. The cerebellum in both age groups contained a distinct neuroepithelial cap and an EGL, which was somewhat better formed in the 12-month-old tadpole. Some heavily labeled cells were found in the neuroepithelial caps of 6-month-old tadpoles from injection times of 6 weeks to 6 months. In the cerebella of 12-month-old tadpoles, however, heavily labeled cells were found in the neuroepithelial cap only with the injection time of 12 months; with injection times from 7 to 11 months, the cells were labeled lightly. Labeled EGL cells were found in the cerebella of 6-month-old tadpoles from an injection time of 6 weeks on; with injection times from 10 weeks to 6 months some EGL cells contained heavy amounts of label. In the cerebella of 12-month-old tadpoles, labeling of EGL cells was not detectable with injection times of 7-9 months; they contained light to medium labeling with injection times of 10 and 11 months and heavy labeling when injected at 12 months. These results indicate that EGL cells are generated continuously in premetamorphic tadpoles from the age of 6 weeks to 12 months. Furthermore, these results suggest that the rate of EGL cell formation is faster during the second half-year of development than during the first.

Animals↗

PubMed1987

Endogenous opioids regulate dendritic growth and spine formation in developing rat brain.

Continuous blockade of endogenous opioid-opioid receptor interaction by opioid antagonists from birth to day 10 increased neuronal maturation in the rat brain. The lengths of oblique dendrites of pyramidal cells in the cerebral cortex and basilar dendrites of the hippocampus were increased from controls by 136 and 51%, respectively, whereas the concentrations of spines in these cells were increased 183 and 69%, respectively. Total dendritic length of spiny branches of cerebellar Purkinje neurons was 65% greater than controls, and spine concentration of granule cells in the dentate gyrus was increased by 76%. Thus, endogenous opioids exert a remarkable influence on the timetable and magnitude of dendritic elaboration and spine formation, and serve as an important trophic influence in the regulation of neuro-ontogeny.

Animals↗

PubMed1987

Androgen action in fetal mouse spinal cord cultures: metabolic and morphologic aspects.

Morphologic and metabolic aspects of androgen effects on the developing spinal cord were studied in organotypic cultures of the E13 (embryonic day 13) fetal mouse lumbosacral spinal cord, maintained as either hemisected, homologous explant pairs co-cultured with bulbocavernosus muscle (morphologic studies), or as whole cross-sectional segments without muscle in which the nutrient medium was supplemented with muscle extract (metabolic studies). Metabolic studies demonstrated the total absence of aromatase activity. 5 alpha-Reductase activity, on the other hand, increased differentially in a segment-dependent manner in spinal cord explants from 0 to 35 days in culture, suggesting regional differences in the utilization of testosterone and its 5 alpha-reduced metabolites. In all studies, spinal cord explants showed androgen-dependent increases in neurite outgrowth, although this was most pronounced in spinal cord-muscle co-cultures. These results indicate that androgens per se affect very early development throughout the entire lumbosacral spinal cord, and that this influence is not restricted to those segments reported to be sexually dimorphic in the adult.

Androgens↗