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Biomedical subjects

I S Zagon

Publications and source records attributed to I S Zagon.

At least 19 recordsLinked to original sources
PubMed1992

Ontogeny of zeta (zeta), the opioid growth factor receptor, in the rat brain.

Opioid growth factor (OGF), [Met5]enkephalin, serves as an inhibitory influence on the developing nervous system and is especially targeted to cell proliferative events. OGF interacts with the zeta (zeta) opioid receptor to perform its function. Using [3H]-[Met5]enkephalin, the ontogeny of the zeta receptor in the whole brain and cerebellum of rats was explored. Specific and saturable binding was recorded at the earliest time sampled, prenatal day 15 (E15). In the whole brain, binding capacity (Bmax) was two-fold greater at E15 than at E18 and E20. The quantity of zeta receptor appeared to increase in the first postnatal week, reaching a maximum on postnatal day 8. Binding decreased the remainder of the 2nd week and between postnatal days 15 and 25 binding was no longer recorded. In the cerebellum, binding capacity increased from E20 to the 2nd postnatal week, reaching a maximum on postnatal days 8-10. The Bmax of the zeta receptor decreased precipitously on postnatal day 11, being 5.4-fold lower than on postnatal day 10. Between postnatal days 21 and 30, no binding was observed. The binding affinities of the whole brain and cerebellum were 2.3 and 2.7 nM, respectively, and no differences between ages could be detected. Continuous opioid receptor blockade from birth to postnatal day 6 increased body weight, the Bmax of the zeta receptor in the whole brain and cerebellum (but not the Kd), and increased the number of layers of germinal cells in the cerebellum.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Developmental expression of brain beta-spectrin isoform messenger RNAs.

We have investigated the expression of brain beta SpIIa and beta SpIb (previously referred to as the beta-subunits of brain spectrin (240/235) and brain spectrin (240/235E), respectively) during mouse brain development. The 9 kb transcript which encodes beta SpIIa is present in fetal mouse brain tissue and increases to a maximal level in a 30-day-old mouse. There is a coordinate accumulation of the 7.8 kb alpha SpIIa mRNA (with beta SpIIa) during mouse brain development. The coordinate expression of alpha SpIIa and beta SpIIa at the mRNA and protein level allows formation of (alpha SpIIa/beta SpIIa)2 tetramers (brain spectrin(240/235)) early in premitotic neuronal development; and avoids turnover of unassembled alpha and beta-subunits. An 11 kb transcript which encodes beta SpIb is not produced in embryonic tissue, and is first seen in a 6-day-old mouse. The protein translation products beta SpIIa and beta SpIb have previously been demonstrated by our laboratory to first appear in fetal mouse brain tissue and at postnatal day 6-8, respectively [J. Neurosci., 7 (1987) 864-874]. The expression of beta SpIb mRNA on postnatal day 6-8, and the appearance of brain spectrin(240/235E) in postmitotic and postmigratory neurons of the cerebellum at this same time; suggests that brain spectrin(240/235E) is involved in differentiated functions of the neuron (formation of cell-cell contacts, formation of dendritic processes and postsynaptic contacts). Thus, the data from the present study demonstrates that the expression of these two neuronal beta-spectrin isoforms is regulated at the level of mRNA expression.

Animals↗
PubMed1992

Identification of an amelin isoform located in axons.

A new axonal isoform of amelin, an analogue of the erythrocyte spectrin binding protein termed protein 4.1 has been identified in mouse brain. This new isoform has a molecular weight of 93 kDa, and migrates to a more acidic pH (pH 7.5-8.0) than the previously described amelin E (pH 8.5) on two dimensional NEPHGE-SDS PAGE. The 93 kDa protein looks nearly identical to amelin E on two dimensional chymotryptic iodopeptide mapping, and both share partial homology with rbc protein 4.1. The new isoform is located in axons, and the soma of neurons in mouse cerebellum, while amelin E is located in neuronal soma and dendrites. The axonal amelin antibody detects a 97 kDa protein in embryonic tissue which diminishes during development; and a 93 kDa protein which is first seen at postnatal day 1 of mouse brain ontogeny, increasing constantly to its adult concentrations. This time course of expression is quite different than amelin E, which is present at embryonic day 15 and diminishes constantly reaching its lowest concentration in the adult brain. We hypothesize that axonal amelin and amelin E may play important roles in the interaction of brain spectrin(240/235) and brain spectrin(240/235E) with f-actin and neuronal membranes.

Aging↗
PubMed1992

An opioid growth factor regulates the replication of microorganisms.

An opioid growth factor (OGF), [Met5]-enkephalin, interacts with the zeta (zeta) opioid receptor to modulate development of eukaryotes. We have found that [Met5]-enkephalin, an endogenous opioid peptide serves to inhibit the growth of S. aureus. This effect on growth involves cell proliferative events and is under tonic control, since potent opioid antagonists accelerate cell replication. Both the OGF and zeta opioid receptor were associated with these microorganisms. Other opioid receptors (mu, delta and kappa) were not detected. OGF also controlled the growth of other bacteria: P. aeruginosa and S. marcesans. These results indicate that OGF and its receptor, known to be important in the regulation of mammalian development, also function in the growth of simple unicellular organisms. We suggest that the endogenous opioid system related to growth originated billions of years ago.

Enkephalin, Methionine↗
PubMed1992

Antinociceptive properties of tiletamine-zolazepam improved by addition of xylazine or butorphanol.

A combination of tiletamine HCl and zolazepam HCl is frequently used as an anesthetic, but little is known about the antinociceptive properties of tiletamine-zolazepam. The antinociceptive properties of tiletamine-zolazepam alone or combined with xylazine or butorphanol were determined in the adult male rate using the tail-flick test. Changes in tail-flick latency were determined at 15, 45, and 75 min after IP drug administration of sterile water, sodium pentobarbital, morphine, tiletamine-zolazepam, xylazine, butorphanol, and tiletamine-zolazepam plus xylazine or butorphanol. Tail-flick latency approximated 100% maximum possible effect (MPE) at 15-75 min postinjection in morphine-treated rats. Tiletamine-zolazepam, xylazine, and butorphanol alone, at any dose utilized, produced less than 50% MPE. However, the combination of tiletamine-zolazepam with butorphanol or xylazine increased tail-flick latency approximately three times greater than tiletamine-zolazepam alone. These results demonstrate that: a) consonant with earlier findings, analgesia and anesthesia are independent states; b) tiletamine-zolazepam is not an effective combination with respect to analgesia; but c) in concert with appropriate drugs, it can exhibit potent antinociceptive properties.

Analgesics↗
PubMed1991

Zeta (zeta), a growth-related opioid receptor in developing rat cerebellum: identification and characterization.

Endogenous opioids and opioid receptors (i.e. endogenous opioid systems) are expressed during neural ontogeny, and play a role in the development of the nervous system. Using [3H][Met5]-enkephalin, a potent ligand involved in neural growth, particularly cell proliferation, specific and saturable binding was detected in homogenates of 6-day-old rat cerebellum; the data were consistent with a single binding site. Scatchard analysis yielded a binding affinity (Kd) of 2.2 nM and a binding capacity (Bmax) of 22.3 fmol/mg protein. Binding was linear with protein concentration, dependent on time, temperature, and pH, and was sensitive to Na+, Mg2+, and guanyl nucleotides. Optimal binding required protease inhibitors, and pretreatment of the homogenates with trypsin markedly reduced binding, suggesting that the binding site was proteinaceous in character. The [Met5]-enkephalin binding site was an integral membrane protein located in the nuclear fraction. Competition experiments indicated that [Met5] enkephalin was the most potent displacer of [3H][Met5]-enkephalin, and that binding was stereospecific. In the adult rat cerebellum, non-opioid receptor binding of [3H][Met5]-enkephalin was recorded, mu and kappa receptors were also found in the developing rat cerebellum, while mu, delta, and kappa receptors were recorded in adult cerebellar tissue. The function, pharmacological and biochemical characteristics, subcellular distribution, and temporal expression of the [Met5]-enkephalin binding site suggest the presence of a unique opioid receptor, termed zeta (zeta), in the developing nervous system.

Animals↗
PubMed1991

Endogenous opioids regulate cell proliferation in the retina of developing rat.

The role of endogenous opioids and opioid receptors (endogenous opioid systems) in modulating cell proliferation in the developing mammalian retina was examined in 1-day-old rats. In contrast to a labeling index (LI) of 35.8% in control animals, administration of the opioid peptide [Met5]-enkephalin (100 micrograms/kg) significantly reduced (10.6%) the proportion of cells incorporating [3H]thymidine; concomitant injection of 1 mg/kg naloxone blocked the inhibitory effects of [Met5]-enkephalin on cell division. Naloxone (1 mg/kg) alone did not alter the LI. The interruption of endogenous opioid-opioid receptor interaction by naltrexone (50 mg/kg), a potent opioid antagonist, was accompanied by a significant increase (6.4%) in the LI relative to control levels. Immunocytochemical experiments revealed the presence of enkephalin-like immunoreactivity, with staining of the cortical cytoplasm of proliferating and differentiating retinal cells recorded; no immunoreactivity was noted in the adult retina. In vitro autoradiography using 125I-[Met5]-enkephalin indicated that [Met5]-enkephalin binding sites were localized to the developing retina; no binding of the radiolabeled ligand was recorded in the adult retina. These results demonstrate the presence of growth-related endogenous opioids and opioid receptors in the developing mammalian retina, but not in adult retina, and suggest that endogenous opioids serve as natural inhibitory trophic factors that tonically regulate cell proliferation.

Animals↗
PubMed1991

Identification of opioid peptides regulating proliferation of neurons and glia in the developing nervous system.

Endogenous opioid systems (i.e. opioids and opioid receptors) play a role in regulating neural development. Using the cerebellar cortex of 6-day-old rats, the most potent opioid peptides involved with cell proliferation were assessed. In both the external germinal (granule) layer (EGL), a germinative matrix giving rise to neurons, and the medullary layer (MED), a pool of cells that are the precursors of glia (astrocytes and oligodendrocytes), [Met5]enkephalin and peptide F were extremely potent in depressing the labeling index (LI) using [3H]thymidine and autoradiographic techniques; concentrations as low as 100 micrograms/kg reduced the LI of EGL cells by 24% and MED cells by 43%. This inhibition of DNA synthesis by opioid peptides was blocked by concomitant exposure to to naloxone, an opioid antagonist. Peptide action was apparent 2 h following drug administration, and concentrations of 80 micrograms/kg but not 1 or 10 micrograms/kg [Met5]enkephalin depressed the LI. These results identify a selective group of opioid peptides, derived from proenkephalin A, as the potent, natural, inhibitory factors targeted to cell proliferation of cells destined to be neurons and glia in the developing nervous system.

Animals↗
PubMed1991

Localization of preproenkephalin A mRNA in the neonatal rat retina.

An opioid growth factor, [Met5]-enkephalin, is known to regulate developmental events in the neonatal rat retina. This growth factor interacts with the zeta (zeta) opioid receptor to modulate retinal ontogeny. Both peptide and receptor are present in developing retina, but not in adult retina. We have used in situ hybridization histochemistry to identify and localize preproenkephalin A mRNA in the neonatal rat retina. Preproenkephalin mRNA was localized to the ganglion cell layer, with some radiolabeling found in the neuroblast layer. This result indicates that 1) the mRNA to preproenkephalin A is present during the critical stage of development in the neonatal retina that coincides with the presence of the growth-regulating peptide, [Met5]-enkephalin, and 2) that the source of the opioid growth factor controlling the production of retinal cells appears to be autocrine (i.e., retinal neuroblasts) and paracrine (i.e., ganglion cells) in nature.

Animals↗
PubMed1991

Synapsin I-mediated interaction of brain spectrin with synaptic vesicles.

We have established a new binding assay in which 125I-labeled synaptic vesicles are incubated with brain spectrin covalently immobilized on cellulosic membranes in a microfiltration apparatus. We obtained saturable, high affinity, salt- (optimum at 50-70 mM NaCl) and pH- (optimum at pH 7.5-7.8) dependent binding. Nonlinear regression analysis of the binding isotherm indicated one site binding with a Kd = 59 micrograms/ml and a maximal binding capacity = 1.9 micrograms vesicle protein per microgram spectrin. The fact that the binding of spectrin was via synapsin was demonstrated in three ways. (a) Binding of synaptic vesicles to immobilized spectrin was eliminated by prior extraction with 1 M KCl. When the peripheral membrane proteins in the 1 M KCl extract were separated by SDS-PAGE, transferred to nitrocellulose paper and incubated with 125I-brain spectrin, 96% of the total radioactivity was associated with five polypeptides of 80, 75, 69, 64, and 40 kD. All five polypeptides reacted with an anti-synapsin I polyclonal antibody, and the 80- and 75-kD polypeptides comigrated with authentic synapsin Ia and synapsin Ib. The 69- and 64-kD polypeptides are either proteolytic fragments of synapsin I or represent synapsin IIa and synapsin IIb. (b) Pure synapsin I was capable of competitively inhibiting the binding of radioiodinated synaptic vesicles to immobilized brain spectrin with a Kl = 46 nM. (c) Fab fragments of anti-synapsin I were capable of inhibiting the binding of radioiodinated synaptic vesicles to immobilized brain spectrin. These three observations clearly establish that synapsin I is a primary receptor for brain spectrin on the cytoplasmic surface of the synaptic vesicle membrane.

Animals↗
PubMed1991

Spectrin isoforms in the mammalian retina.

Spectrin is a major component of the mammalian neuronal cytoskeleton. In the CNS, three isoforms of brain spectrin are known to exist: a cellular and dendritic isoform, (240/235E), related to neurons and glia; a cellular and axonal isoform, (240/235), related to neurons; and an isoform specific for astrocytes, (240/235A). In the present study, brain spectrins (240/235E) and (240/235) were localized within the mouse retina and optic nerve. Immunoblot analyses of proteins isolated from mouse retinas utilizing polyclonal antibodies to either brain spectrin (240/235) or brain spectrin (240/235E) revealed that these spectrins are present in the retina and that the two isoforms are the same molecular weights as those found in the brain. Immunocytochemical studies revealed that spectrin (240/235E) was localized in cell bodies of the inner nuclear, outer nuclear, and ganglion cell layers, and processes arborizing within the inner and outer plexiform layers. Spectrin (240/235) was distributed diffusely within the retina, lightly staining neurons in both the inner nuclear and outer nuclear layers, and the ganglion cell layer. In contrast to the situation found in the brain, spectrin (240/235) was but one of the axonal forms in the retina. We found that spectrin (240/235E) was also present in the axon-rich fiber layer and in the optic nerve and was often associated with fibrous elements. Spectrin (240/235) was also detected in the nerve fiber layer and optic nerve, but this isoform was not localized to fibers.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Adult and developing human cerebella exhibit different profiles of opioid binding sites.

The binding of [3H][D-Ala2,MePhe4,Gly-ol5]enkephalin (DAGO), [3H][D-Pen2,5]enkephalin (DPDPE), [3H]ethylketocyclazocine (EKC), and [3H][Met5]enkephalin (MET) was used to examine mu-, delta-, kappa-, and zeta-receptors, respectively, in the developing (birth to postnatal day 19) and adult human cerebellum. Specific and saturable binding of all ligands was recorded in developing brains, and of [3H]DAGO, [3H]DPDPE, and [3H]EKC in adult cerebellum; all data fit a single homogeneous binding site for each ligand. However, the ontogenic profile of opioid receptor subtypes differed. Delta- and kappa-receptor capacities were 7.8- and 3.6-fold, respectively, greater in infant cerebellum than in adults. The mu-receptor decreased over 7-fold in both binding affinity and capacity after day 2; by adulthood, the binding affinity was the same as in newborns but only one-half the binding capacity was recorded. The concentration of zeta-receptors was 20-fold greater in subjects 2-19 days of age than in newborns. These data demonstrate the presence, and distinct developmental profiles, of opioid receptors in human cerebellum. Although the function of mu-, delta-, and kappa-receptors in human cerebellum are unclear, the growth-related zeta-receptor is present at a time of cell replication and differentiation but is not detected in mature cerebellum.

Adult↗
PubMed1990

Demonstration and characterization of zeta (zeta), a growth-related opioid receptor, in a neuroblastoma cell line.

Endogenous opioids and opioid receptors (i.e. endogenous opioid systems) are involved in carcinogenesis. Using homogenates of S20Y neuroblastoma (NB) cells grown in culture, the binding of a growth-selective ligand, [Met5]enkephalin, was examined to ascertain the zeta (zeta) opioid receptor. Specific and saturable binding of [3H]-[Met5]enkephalin was detected in NB cells; the data were consistent with a single binding site. Scatchard analysis yielded a Kd of 1.6 nM and a binding capacity (Bmax) of 48.1 fmol/mg protein; 14,000 receptors per cell were estimated. Binding was dependent on protein concentration, time, temperature, and pH, and was sensitive to 100 nM, but not 5 nM, Na+, Ca2+, and Mg2+; GppNHp at concentrations of 100-500 mM had little effect on binding. Optimal binding required protease inhibitors, and pretreatment of the tumor cell homogenates with trypsin markedly reduced [3H]-[Met5]enkephalin binding, suggesting that the binding site was proteinaceous in character. Displacement experiments indicated that [Met5]enkephalin was the most potent displacer of [3H]-[Met5]enkephalin. Cell density (log, confluence, postconfluence) did not alter the Kd or Bmax. This study serves as the first demonstration and characterization of the zeta (zeta) opioid receptor in tissue culture cells. The homogeneous nature of NB cell cultures, along with the enrichment in receptor number, provides an excellent model system to isolate and purify the zeta receptor.

Animals↗
PubMed1979

Morphological identification and biochemical characterization of isolated brain cell nuclei from the developing rat cerebellum.

Cell nuclei from developing rat cerebellum were isolated and the various types of nuclei were characterized and quantified. Nuclear pellets appeared to be both quantitatively and qualitatively representative of the entire cerebellum, and of sufficient purity to perform biochemical studies as well as morphological comparison with histological sections. Isolated nuclei were classified into 6 groups based on nuclear size and shape, heterochromatin aggregations, and nucleoplasmic density. The total population of cerebellar cells primarily consisted of two types of nuclei after day 10. One group of nuclei, resembling those of internal granule neurons or external germinal cells, contributed at least 70% of the total isolated cell nuclei from day 1 to day 90, whereas another nuclear group that was identified as dark oligodendrocytes constituted 8-9% of the total population on days 45 and 90. Nuclear DNA, RNA, and protein content of the cerebellum also were determined throughout postnatal development. DNA concentration markedly declined after day 15, while the RNA/DNA ratio increased until day 3 and remained constant to day 90. The nuclear protein/DNA ratio increased from birth to day 3, decreased to its lowest value on day 10, and increased to day 90. Utilizing DNA values, the total cell population as well as contributions of different cell types were calculated. At birth the cerebellum was estimated to contain 5.9 million cells, increasing to 94 million by day 21. By day 90, 107 million cells were present, of which 8.6 million oligodendrocytes and 93.6 million internal granule cells were estimated.

Aging↗
PubMed1979

Hypophagia follows the initial hyperphagia produced by 2-deoxy-D-glucose in rats.

The glucose analogue 2-deoxy-D-glucose (2DG) inhibits glucose metabolism and causes a rapid increase in food consumption in most species. This increase is most apparent during the first 6 postinjection hours, although it may persist as long as 10 hr. There are no published descriptions of alterations in food consumption subsequent to the hyperphagia. In the present study male and female rats were injected with 2DG (750 mg/kg IP), insulin (regular, 20 U/kg SC) or distilled water, and food intake was compared to baseline levels during the next 1, 6 and 24 hr. Results showed that food intake: (1) was not affected by injections of water: (2) was higher than normal during all 3 time periods following insulin injections: and (3) was higher than normal at 1 and 6 hr following 2DG, but significantly lower than normal by the end of 24 hr. The reasons underlying the development of hypophagia subsequent to the initial hyperphagia produced by 2DG are presently unknown.

Animals↗
PubMed1979

Learning ability in adult female rats perinatally exposed to methadone.

Cognitive functioning of adult female rats that were maternally exposed to methadone (5 mg/kg daily) during gestation and/or lactation was studied by assessing performance on a food-motivated light-dark discrimination learning test and on active and passive shock-avoidance tests. Methadone-exposed rats exhibited difficulties on the light-dark discrimination learning and the active avoidance tests, and behavioral deficits appeared to be related to the timing and duration of drug treatment. On the light-dark discrimination test only 33% of the rats in the gestation group and 25% of the animals in the lactation group met criterion in comparison to 87% of the control rats. Thirty-three percent of the animals in either the gestation or gestation-lactation groups met criterion on the active avoidance test in contrast to 87% of the controls. These data suggest that perinatal exposure to methadone impairs cognitive abilities in the adult female rat.

Animals↗
PubMed1979

Imparied thermal regulation in juvenile rats following perinatal methadone exposure.

Offspring of female rats injected daily with methadone (5 mg/kg) or saline were cross-fostered at birth to form groups exposed to methadone during gestation (G), lactation (L), or gestation and lactation (G-L); controls (C) were exposed only to saline. Rectal temperature, body weight and food consumption were measured from postnatal Days 36-51. Ambient temperature was maintained at 21 degrees C except for Days 42--45, when the temperature was 10 degrees C. Group G rats never differed from controls, but offspring in Groups L and G-L were hypothermic at room temperature; Group G-L rats exhibited a further temperature loss during the cold stress. There were no group differences in food consumption after Day 39, and all groups increased food intake while in the cold. Group differences in body weight were not reliable but Group G-L rats gained less weight than the rest during the experiment, whereas Group L rats gained more. These results indicate that, depending upon treatment schedule, perinatal methadone exposure is associated with hypothermia during the postweaning period. A prolonged withdrawal reaction from methadone may account for the impaired thermal regulation.

Animals↗