Receptors In Rat Liver Research Articles

Modification of arginine residues in human growth hormone by 1,2-cyclohexanedione: Effects on the binding capacity to lactogenic and somatogenic receptors

Reactivity of arginine residues in human growth hormone was studied by reaction with 1,2-cyclohexanedione. Kinetic analysis of the data showed a good fit to a pseudo first order curve, with an apparent velocity constant k = 1.26 × 10 −2 min −1 and a maximum modification of 9.6 out of the 11 arginines of the molecule. Modification led to a decrease in binding capacity to both lactogenic and somatogenic rat liver receptors. In either case Tsou plots suggest that the modification of two arginine residues is responsible for this behavior, although it cannot be ascertained whether the two relevant residues are the same for both receptor types. Circular dichroism studies indicated no apparent changes in protein conformation in the modified hormone. Binding capacity was restored upon regeneration of arginines by incubation with Tris-HCl buffer. Only the carboxy-terminal peptide was isolated by HPLC from a tryptic digest of succinylated Arg-modified hGH, indicating that 183 is the nonreacting arginine residue.

Effects of chronic ethanol intake on aromatization of androgens and concentration of estrogen and androgen receptors in rat liver

The present study was carried out to investigate if ethanol alters aromatization of androgens and concentrations of hepatic estrogen and androgen receptors. Hepatic aromatization of androgen to estrogen was significantly increased by ethanol administration. There was a significant increase in serum estrogen level but a decreased circulating testosterone level in alcohol-fed rats. Furthermore, the concentration of estrogen receptors in liver cytosol was significantly higher in alcohol-fed rats (37 ± 5.3 fmol/mg protein), as compared to the intact control value (21 ± 4.8 fmol/mg protein). However, hepatic androgen receptor levels were much lower (4.4 ± 0.5) in alcohol-fed rats than those (10.2 ± 1.4 fmol/mg protein) in control animals. Similarly, castration increased hepatic aromatization of androgens and concentrations of serum estrogen and hepatic estrogen receptors, but it decreased contents of circulating androgen and hepatic androgen receptors. These findings indicate that alcohol administration is considered a chemical form of castration, altering the hepatic steroid metabolism and sex hormone receptor contents and contributing to the pathogenesis of ferminization. A combination of alcohol-feeding and castration has no synergistic effect on the hepatic steroid receptors and aromatization, but this combination does have a more profound effect in lowering the concentration of circulating androgen.

Isolation and characterization of two novel rat ovarian lactogen receptor cDNA species

Two novel lactogen receptor cDNA clones (2.1 and 1.2 kb) were isolated from a rat ovarian cDNA library. Nucleotide sequence of the 2.1 kb clone codes for a 610 aa receptor (nonglycosylated mol. wgt. 66,000 D) with an extracellular domain, a transmembrane region and an intracellular domain, and exhibited significant overall similarity with the rat liver receptor (310 aa) and both rabbit mammary and human hepatoma receptors (616 and 622 aa). However, the ovarian lactogen receptor sequence contains a unique cytoplasmic domain of 110 aa and consensus sequences for both a tyrosine phosphorylation site and an ATP/GTP type A binding site, and thus has potential for signal transduction and mitogenic activity. The 1.2 kb clone codes for a truncated binding form of 150 aa that is identical with the ovarian long form over only the first 130 residues, and lacks the transmembrane region. Differences between long and short forms of the ovarian lactogen receptors and the truncated liver species may result from alternative splicing. The prolactin holoreceptor gene(s) has the potential for producing several receptor subtypes that differ in tissue-specific expression, size, compartmentalization and mode of signal transduction, and may subserve the divergent functions of prolactin in its several target cells.

Insulin-like growth factor I receptors in adult rat liver: characterization and in vivo regulation

Although the presence of significant amounts of insulin-like growth factor (IGF)-I receptors in fetal tissues is well documented, adult liver has been reported to contain little or no IGF-I binding activity. In the present investigation, substantial amounts of specific IGF-I receptors were detected in crude membrane fractions and in partially purified receptor preparations of female adult rat liver. Insulin was 100 times less potent than IGF-I in competing for 125I-IGF-I binding. IGF-I binding activity was much less than that of insulin binding in both the microsomal fraction and partially purified receptor preparations. Affinity cross-linking of 125I-IGF-I to purified receptors and microsomal fractions followed by electrophoretic analysis under nonreducing conditions revealed labeling of proteins with relative molecular weight (Mr) of 350,000 and 210,000-220,000, corresponding to the molecular mass of the intact tetramer and alpha-beta dimers, respectively. Under reducing conditions, the labeling of proteins with Mr of 130,000 and 260,000, corresponding to the alpha-subunit of IGF-I receptor and its dimer, respectively, was observed. Treatment of microsomes as well as partially purified receptors with 0.5-1 mM dithiothreitol resulted in decreased IGF-I binding, and this correlated with structural changes in the receptor as detected by affinity labeling and electrophoretic analysis. Hepatic IGF-I binding activity was significantly diminished in female rats exposed to chronic growth hormone excess, suggesting down-regulation of IGF-I receptors by the enhanced circulating levels of IGF-I in these animals.(ABSTRACT TRUNCATED AT 250 WORDS)

Characterizations of two forms of inositol 1,4,5-trisphosphate receptor in rat liver

Two binding sites for [ 32P]myo-inositol 1,4,5-triphosphate (Ins(1,4,5)P 3) were detected in a crude particulate fraction prepared from rat liver homogenate and in permeabilized hepatocytes. The same high- and low-affinity sites with K D s of 1.8–2.6 nM and 35–71 nM, respectively, were detected in subcellular fractions enriched in plasma membranes, mitochondria and microsomes, with relative proportions close to those found in the crude membrane function. The order of potency of three inositol phosphates in inhibiting [ 32P]Ins(1,4,5)P 3 binding to the two sites, i.e. Ins(1,4,5)P s > Ins (2,4,5(P 3] > Ins(1,3,4,5)P 4, and the inhibition by heparin, strongly suggest that neither of the binding sites reflected components due to the 3-kinase or the 5-phosphatase. A close correlation was observed between the dose-response curves for Ca 2+ release by Ins(1,4,5)P 3 and Ins(2,4,5)P 3 and the occupancy of the low-affinity binding site by these agonists. These results support the view that the two [ 32P]Ins(1,4,5)P 3 binding sites are two forms of the same receptor.

Heterogeneity of rat ovarian lactogen receptor species

Three distinct ovarian lactogen receptor species with unique and highly reproducible HPLC retention times gave corresponding peaks of binding activity and migrated as single bands of 80, 40, and 34 kDa on SDS-PAGE. Reduction of the 80 kDa protein failed to reveal any conversion to the lower molecular weight proteins by either SDS-PAGE analysis or reverse phase HPLC, suggesting that the three binding proteins are not related by disulfide bond formation. Immunological studies indicate an amino acid homology at a C terminal region among the 3 receptor forms and with the rat liver receptor. However, the 80kDa ovarian receptor also contains a unique sequence derived from microsequencing that is not immunologically apparent in the ovarian lower molecular weight forms of the rat liver receptor. These findings substantiate the existence of at least two populations of ovarian lactogen receptors perhaps originating within the same gene, a high Mr form potentially capable of signal transduction, and truncated forms that could be involved in transport and/or clearance.

Stable association of solubilized VIP receptor and Gs protein in rat liver: Immunological and functional evidences

Stable association of solubilized VIP receptor and Gs protein in rat liver: Immunological and functional evidences

Immunohistochemical demonstration of the asialoglycoprotein receptor in rat liver by a sensitive avidin-biotin technique.

An immunohistochemical technique for the detection of the hepatic asialoglycoprotein receptor (ASGP-R) in cryostat sections of liver by polyclonal and monoclonal anti-ASGP-R antibodies is described. The procedure is based on the alkaline-phosphatase-avidin-biotin complex (ABC-AP) system and important features include fixation of the sections with periodate-lysine-paraformaldehyde (with or without dichromate) and an absolute requirement for blocking of endogenous biotin activity. The sensitivity of the technique is such that binding to ASGP-R can be detected with femtomolar concentrations of monoclonal anti-ASGP-R antibodies and, with polyclonal antisera, approaches that of a radioimmunoassay.

Regulation of beta‐adrenergic receptor expression in rat liver

To begin defining the factors regulating neurotransmitter receptor expression, we examined beta-adrenergic receptors in rat liver in vivo and in primary liver cultures under defined hormonal conditions. beta-receptors described a remarkable developmental profile in vivo, increasing fivefold between embryonic days 16 and 20, and decreasing tenfold by early adulthood. The developmental decrease reflected reduced receptor number without a change in receptor properties. The ontogenetic decrease appeared to be specific for beta-receptors; alpha-receptors developed in a hyperbolic fashion, reaching high plateau values by the third postnatal week. Adult rat liver cells plated into culture re-expressed high beta-receptor levels, exhibiting a 4-8-fold increase. A similar pattern of expression of the beta-receptors, having similar pharmacological properties, was observed in primary liver cultures maintained in serum-free medium, in a serum-supplemented medium or in several variations of a serum-free, hormonally defined medium designed for primary liver cultures. Thus, the degree of expression of the beta-receptors was not found affected by various hormones, by serum, or by any medium condition. By contrast, the degree of expression of the beta-receptors was markedly sensitive to cell density. High expression of the beta-receptors was observed at low cell densities (1-3 x 10(6) cells/150 mm dish), and low expression or no expression was observed in confluent cultures (10-20 x 10(6) cells/150 mm dish). Our experiments suggest that beta-receptor expression does not follow an immutable program, but may be regulated by density-dependent cell-cell interactions.

Chimeric bovine-human growth hormone prepared by recombinant DNA technology: binding properties and biological activity.

A chimeric bovine GH (amino acids Met-Asp-Gln-greater than 1-23) and human GH (hGH) (amino acids 24-191) plasmid was constructed and expressed in Escherichia coli. The purified protein (chimeric GH) exhibited a 2-3 order of magnitude lower affinity toward lactogenic receptors in Nb2 lymphoma cells, microsomal fractions from bovine mammary gland and male rat liver. The affinity towards somatogenic receptors in IM-9 human lymphocytes and male rat liver was decreased to a much lesser degree. This diminished affinity towards lactogenic receptors was accompanied by a parallel decrease in the ability of the chimeric GH to stimulate proliferation of Nb2-11C lymphoma cells and the lipogenesis in bovine mammary gland. This implies that occupation of the respective receptors by either chimeric GH or hGH leads to identical postreceptoral effects. The chimeric GH was also capable of down-regulating the lactogenic receptors in Nb2 lymphoma cells and was recognized by three anti-hGH monoclonal antibodies. These and previously published results indicate that the N-terminal part of hGH is essential for the high affinity binding to lactogenic receptors and subsequent biological effect. Removal or replacement by a corresponding part of bovine GH converts the hormone, respectively to weak antagonist or agonists. Analysis of our data, based on hydropathy index leads us to suggest that the high affinity binding site of the hGH towards lactogenic receptors is mainly confined to amino acids nos. 8-18.

Immunofluorescent localization of type II insulin-like growth factor receptor in rat liver and hepatoma cells

We have used an immunofluorescent technique to localize type II insulin-like growth factor (IGF) receptors in rat liver, rat hepatocytes and three rat hepatoma cell lines (HTC, H-35 and 5123) using a polyclonal antibody (C-1) raised to purified rat liver type II IGF receptor. Specificity of the antiserum was confirmed by Western blotting of microsomal membranes prepared from hepatocytes and hepatoma cells which showed a single class of receptor in all cells, of Mr approximately 210,000 for hepatocytes, HTC and H-35 cells and approximately 220,000 for 5123 cells, on non-reduced, 4-15% polyacrylamide gradient gels. The specificity of the immunofluorescent technique was also verified by abolition of labelling after preincubation of antiserum with purified type II IGF receptor. Rat liver cryosections contained areas of juxtanuclear labelling in hepatocytes, consistent with the presence of type II IGF receptor in the Golgi region. Brightest immunofluorescence was seen in sections from fetal and neonatal rats with adult rat hepatocytes staining brightly only around central veins. Areas of labelling were also seen in connective tissue surrounding larger veins. Cultured adult rat hepatocytes and rat hepatoma cell lines also showed bright areas of juxtanuclear nuclear immunofluorescence, with HTC and H-35 cells staining more than 5123 and adult hepatocytes. Fetal rat hepatocytes in culture also labelled very brightly both in a juxtanuclear location and in small clusters over the cell, possibly on the cell surface. These observations indicate that type II IGF receptors are located predominantly on intracellular membranes and are most abundant in rapidly growing cells and tissues (such as fetal liver and hepatoma cells).

Characterization and applications of monoclonal antibodies to the prolactin receptor.

Monoclonal antibodies (mAbs) were produced in BALB/c mice immunized with partially purified PRL receptors from rat liver. Two mAbs (T1 and T6) were able to completely inhibit [125I]ovine PRL ([125I]oPRL) binding to solubilized rat liver PRL receptors, while two other mAbs (U5 and U6) showed only a small effect on PRL binding, but were able to precipitate hormone-receptor complexes. Scatchard analysis of [125I]oPRL binding to rat liver microsomes in the presence of mAbs resulted in a decrease in the number of sites without changing the affinity of PRL binding by T1 and T6, whereas U5 and U6 altered neither parameter. [125I]mAb binding to rat liver microsomes was performed in the presence of various concentrations of unlabeled mAbs or oPRL to examine the interaction between mAbs. Competition of binding to the receptor was observed, respectively, between T1 and T6, U5 and U6, and U5 and E21 (a mAb to the rat liver PRL receptor previously produced). Both [125I]T1 and [125I]T6 binding were inhibited by oPRL, although not completely (80% inhibition at the higher concentrations). When [125I]T1 binding was analyzed by Scatchard analysis, two classes of binding sites to rat liver microsomes were found, of which only the number of higher affinity sites was affected by the presence of oPRL in incubation. Similar results were observed for [125I]T6 binding. [125I]mAb binding to microsomes from other tissues and species was examined. All five mAbs were able to bind to microsomes from rat tissues (liver, ovary, adrenal, prostate, and Nb2 lymphoma cells), similar to the level of [125I]oPRL binding in these tissues. The binding characteristics of [125I]T6 or [125I]U5 were essentially identical in all rat tissues examined. Although T1, U6, and E21 showed strong species specificity, there was significant binding of T6 to rabbit liver and mammary gland and of U5 to rabbit and pig mammary gland and mouse liver Competition curves of [125I]U5 binding were parallel for rat, rabbit, and mouse tissues, while [125I]T6 binding was able to distinguish PRL receptors in rabbit mammary gland from those in rat tissues. The use of 125I-labeled mAb in immunoblot analysis of the PRL receptor resulted in a marked increase in sensitivity. All mAbs detected microsomal PRL receptors in rat liver with mol wt of 84,000, 42,000 and 40,000. As little as 4 fmol receptors can be identified using this approach. Microsomal PRL receptors from rat ovary, prostate, and Nb2 cells and purified receptors from pig and rabbit mammary gland were subjected to immunoblot analysis.(ABSTRACT TRUNCATED AT 400 WORDS)

Hormonal Regulation of Inositol 1,4,5-trisphosphate Receptor in Rat Liver

Inositol 1,4,5-trisphosphate (IP3) is a second messenger which induces Ca2+ release from an intracellular store. We have investigated the properties of the [32P]IP3 binding sites in rat liver. Two specific [32P]IP3 receptors with KD of 2.3 and 88 nM and respective capacities of 33 fmol/mg protein and 195 fmol/mg protein have been detected in a crude membrane fraction prepared from rat liver homogenate. The pretreatment of the liver with IP3-dependent hormones increased two-fold the capacity of the high affinity site. This effect was partly reversed by dibutyryl cyclic AMP. Permeabilized hepatocytes also displayed two [32P]IP3 binding sites with KD of 1.5 and 84 nM and respective capacities of 8 and 300 fmol/10(6) cells. We have measured the [32P]IP3 binding and the IP3-induced 45Ca2+ release in the same batch of permeabilized hepatocytes. In a low Mg2+ medium, the EC50 for 45Ca2+ release was in close correlation with the KD for the low affinity site. These data suggest that an equilibrium between two states of the IP3 receptor is regulated by hormone action and the low affinity state is responsible for the intracellular Ca2+ release.

Nitrosylated high density lipoprotein is recognized by a scavenger receptor in rat liver.

In order to assess the presence of specific recognition sites for high density lipoprotein (HDL) in vivo, HDL was nitrosylated with tetranitromethane and the decay and liver uptake were compared with that of native HDL. The association of intravenously injected nitrosylated HDL (TNM-HDL) with liver was greatly increased as compared to native HDL. Using a cold cell isolation method, it became evident that the liver endothelial cells were responsible for the increased uptake of the modified HDL. The involvement of the endothelial cells in the uptake of TNM-HDL from the circulation could also be demonstrated morphologically by using the fluorescent dye dioctadecyl-tetramethyl-indocarbocyanine perchlorate (Dil) to label HDL. In vitro competition studies with isolated liver endothelial cells indicated that unlabeled modified HDL and acetylated LDL displaced iodine-labeled TNM-HDL, while no competition was seen with LDL and a slight displacement was seen with unlabeled native HDL. Nonlipoprotein competitors of the scavenger receptor such as fucoidin and polyinosinic acid blocked the interaction of TNM-HDL with the liver endothelial cells. Also the degradation of TNM-HDL was blocked by low concentrations of chloroquine. It can be concluded that a scavenger receptor on liver endothelial cells is involved in the clearance of tetranitromethane-modified HDL, which excludes the possibility of using TNM-HDL in vivo to assess the non-receptor-dependent uptake of HDL. The use of nitrosylated HDL in vitro as a low affinity control is limited to cell types that do not possess scavenger receptors, because cell types with scavenger receptors will recognize and internalize TNM-HDL by a high affinity scavenger pathway.

Differences in nuclear thyroid hormone receptors among species

Hepatic nuclear thyroid hormone receptors from rat, dog, chicken, and rainbow trout were compared. Receptor affinities for 3,5,3′-triiodo- l-thyronine (T 3) were similar in preparations from rat, dog, and chicken, using isolated nuclei and nuclear extracts. Rainbow trout nuclear receptor showed a lower affinity for T 3. Almost half of the receptors were released into the medium with rat and chicken nuclei, and 79.7 ± 1.1% of the receptors were released with rainbow trout nuclei, when isolated nuclei were incubated with T 3 at 22° for 2 hr. The affinity constant of rat liver receptor for calf thymus DNA-cellulose at 0.17 M KCl, pH 7.4, was 3.98 ± 1.47 × 10 5 M −1, when determined using DNA-cellulose columns. The number of salt bridges involved in DNA binding of the rat receptor was 5.73 ± 0.38. When receptor-DNA interactions were compared among species, significant differences were found, but the receptors from dog and rainbow trout liver were similar. Sephacryl S-200 column chromatography showed that chicken receptor had a Stokes radius significantly smaller than that of rat receptor. Partial proteolysis of T 3-receptor complex using trypsin α-chymotrypsin, elastase, and papain produced distinct T 3-binding fragments in different species. Our data provide evidence that nuclear thyroid hormone receptors from different species have significant structural dissimilarities.

Biogenesis of the Somatogenic Receptor in Rat Liver

Certain structural characteristics, in particular the type of oligosaccharide chains associated with the rat liver somatogenic (GH) receptors, were studied in different isolated organelles involved in receptor biosynthesis, maturation, and binding, with the use of ligand-affinity cross-linking, incubation with various oligosaccharide chain-cleaving enzymes, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In an endoplasmic reticulum-enriched fraction, a somatogenic receptor with Mr 33,000, after correction for bound ligand (assuming a 1:1 binding ratio of ligand to receptor) was found to contain N-linked high mannose oligosaccharide chain(s). In an intermediate density fraction, enriched in cis-Golgi, a major receptor of Mr 43,000 was found to contain N-linked complex type of oligosaccharide chains. In a low density membrane fraction, containing trans-Golgi complex membranes and endocytic vesicles, three receptors of Mr 95,000, 55,000, and 43,000 were found. These three receptors contain N-linked complex-type oligosaccharide chains. Neuraminidase treatment resulted in a decrease of the Mr 95,000 and 43,000 receptors to Mr 81,000 and 39,000, respectively. Two specific somatogenic receptors of Mr 95,000 and 43,000 containing N-linked complex type of oligosaccharides were found in an isolated plasma membrane-enriched fraction. When isolated hepatocytes were analyzed, the Mr 95,000 receptor was found to be the major labeled species. Two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis (first dimension nonreducing and the second dimension reducing conditions), showed that the Mr 43,000 receptor is contained within the Mr 95,000 receptor. The data suggest that the Mr 33,000 receptor found in endoplasmic reticulum constitutes a precursor to the Mr 43,000 receptor and that the Mr 43,000 receptor is complexed with an unknown subunit during transport through the Golgi complex to form an Mr 95,000 receptor present on the cell surface.

Androgen receptor in rat liver: Characterization and separation from a male-specific estrogen-binding protein

Many liver processes are sexually dimorphic. In particular, the microsomal content of specific enzymes and the synthesis of specific proteins are under sex steroid hormone control. Because the liver of male rats is strikingly androgen responsive, we sought evidence for an androgen receptor in this tissue. We detected and characterized both cytosolic and nuclear androgen-binding proteins. Both forms bind [ 3H]R1881 (methyltrienolone, 17β-hydroxy-17α-methyl-4,9,11-estratriene-3-one) with the high affinity, low capacity, and specificity for androgens and antiandrogens characteristic of androgen receptors. No high-affinity binding of [ 3H]DHT could be detected in unfractionated cytosol because of the rapid metabolism of this ligand; however, binding of a DHT metabolite to the high-capacity male-specific estrogen binder (MEB) of cytosol was observed. Both gel filtration and heparin-Sepharose affinity chromatography separate the cytosolic androgen receptor from MEB. Incubation of cytosol in the absence of sodium molybdate resulted in androgen-binding activity which was retained by DNA-cellulose. Castration of male rats results in a time-dependent loss of both cytosolic and nuclear androgen binding, as well as a loss in MEB activity. Androgen-binding activity is low in livers from female rats, but can be induced by testosterone treatment. An intact pituitary is necessary for maintenance of androgen-binding activity, as hypophysectomy results in complete loss of activity.

Effect of forced-running stress on β-adrenergic receptors in rat brain and liver

Effect of forced-running stress on β-adrenergic receptors in rat brain and liver

Insulin-like Growth Factor (IGF) Receptor in Human Fetal Erythrocytes and Fetal Rat Liver

In order to clarify the potential role of IGF-I in fetal growth, the dynamics of IGF receptor were investigated in comparison with insulin receptor in human and rat fetuses. In humans, the serum levels of IGF-I measured by IGF-I radioimmunoassay after acid-ethanol extraction were significantly lower in fetuses than in adult controls. Conversely, serum insulin levels in fetuses were not indistinguishable from those in adults. On the other hand, specific binding of 125I-IGF-I to human fetal erythrocytes was 2.3%, which was significantly higher than that of adult women (1.6%). Insulin binding to the erythrocytes was also higher in human fetuses than in adult controls (cord: 3.8%, adult: 3.0%). By Scatchard analysis, the changes in the specific binding of IGF-I and insulin were mainly due to the changes in the binding capacity rather than those in the binding affinity. Additionally, IGF receptor on human fetal erythrocytes was recognized as type I IGF receptor, because the binding of 125I-labelled IGF-I and 125I-labelled IGF-II could be displaced by the addition of cold IGF-I, IGF-II and insulin. In rats, serum levels of IGF-I were also much lower in fetuses than in adult controls. Specific binding of 125I-IGF-I to liver microsomal membranes was 34.3% (per 400 micrograms protein) in fetal rats (D20), which was significantly higher than that of adult rats (3.2%). Scatchard analysis indicated that these changes in 125I-IGF-I binding was chiefly due to the changes in binding capacity. On the other hand, serum insulin levels increased with progress of pregnancy, and specific binding of insulin to microsomal membranes of fetal liver increased on D21 of gestation, mainly due to the increase in binding affinity rather than binding capacity. The bindings of 125I-labelled IGF-I and 125I-labelled IGF-II to fetal liver microsomal membranes were clearly displaced by cold IGF-I and IGF-II but not by excess of cold insulin, indicating that the receptor was type 2 IGF receptor. These results suggest that IGF-I possesses much more receptor in fetal tissues than in adult tissues despite its lower concentration in fetal sera. It is speculated that IGF-I may play an important role in fetal growth and development.

The heterogeneity of the developing brain insulin receptor.

Comparison of the adult brain insulin receptor (IR) to other tissue IR demonstrates that the former migrates approximately 10 kD faster on sodium dodecyl sulfate-polyacrylamide gel electrophoresis due to deficient sialic acid content of the asparagine N-linked carbohydrate moieties. We studied these receptors in the fetal rat (18-day) brain (approximately 125 kD) and liver (approximately 135 kD), and demonstrated that similar differences are present during fetal life. These differences are not modified by hyperglycemia associated with both mild hyperinsulinemia and normoinsulinemia/hypoinsulinemia. We further studied the specific brain cell types: neurons, glial cells, and purified microvessel preparation, and demonstrated a heterogeneity in the N-linked glycosylation of the IR within an organ (brain). The neuronal (approximately 125 kD) and microvascular (approximately 125 kD, approximately 135 kD) IR are deficient in sialic acid, thus conferring neuraminidase-insensitivity to the whole brain, whereas the glial cell IR, similar to the liver IR, exhibits neuraminidase sensitivity and migrates intermediate (approximately 128 kD) to the liver and brain IR. The functional significance of this receptor heterogeneity between various tissues and cells within the same organ (brain) remains to be determined.