Karyometry and histometry of renal-cell carcinoma

In an attempt to more objectively predict the outcome of renal cancers, karyometric and histometric studies were performed using an interactive computer-based system for the quantitative analysis of tissue sections. Analysis showed a significant relationship between patient survival and metastases and the histometric parameters of nuclear elongation, nuclear crowding and mitotic density, as well as tumor grade. Patients who died tended to have a high mitotic density, elongated and crowded nuclei and high-grade tumors. Ploidy showed no significant correlation with prognosis while nuclear elongation and crowding did. Differences in histologic grade were significantly associated with several histometric variables, including nuclear area, shape, crowding, elongation and mitotic density.     

Subunit structure and higher order assembly of the hemocyanins of the Melongenidae family: Melongena corona (Gmelin), Busycon canaliculatum (Linné), B. carica (Gmelin), B. contrarium (Conrad), and B. spiratum (Lamarck)

1. The hemocyanins of the Melongenidae family of marine gastropods: Melongena corona, Busycon canaliculatum, B. carica, B. contrarium, and B. spiratum exist in solution as multi-decameric aggregates characterized by sedimentation coefficients of approximately 105 S, 130 S, 150 S, 170 S, and higher values, corresponding to di-, tri-, tetra-, penta-, and larger multi-decameric particles. 2. The hemocyanins of B. contrarium and B. carica seem to form the largest decameric aggregates with the tri- to penta-decamers respresenting the major constitutents. Scanning transmission electron microscopy (STEM), both of unstained, freeze-dried and negatively-stained specimens, shows the presence of discrete aggregates consisting of up to ten decameric units. 3. The particle masses as determined by STEM mass measurements for individual molecules gave integral multiples of from 4.2 x 10(6) to 4.4 x 10(6) daltons ranging from about 8.2 x 10(6) daltons for the typical di-decamer of B. canaliculatum hemocyanin to as high as about 39 x 10(6) and 43 x 10(6) for the nano-and deca-decamers of B. contrarium hemocyanin. 4. The appearance of the higher multi-decamers in both negatively-stained and freeze-dried specimens suggest that they are formed by the addition of decameric units to a single di-decameric unit "tail-wise" in both directions. The higher aggregates formed seem to terminate with a closed head or collar at both ends of the assembly.     

Adrenergic interactions in uterus and vascular smooth muscle in rats in vivo

Simultaneous blood pressure and uterine responses to norepinephrine infusions were recorded in urethane-anesthetized, pentolinium-indomethacin treated rats in natural estrus under conditions in which no blockers or blockers of alpha 1-, alpha 2-, and beta-adrenergic receptors or of "reuptake" of norepinephrine were present. The contributions of alpha 1- and alpha 2-adrenergic receptors to the blood pressure response were similar during the initial portion of the response. At later times, however, alpha 1-adrenergic receptors were responsible for the major portion of the response. The tachyphylaxis of the pressor response that occurs during norepinephrine infusion could be prevented by preventing norepinephrine "reuptake" with imipramine. In the uterus, the initial small alpha-adrenergic contractile response (seen only at the lowest infusion rate) was quickly overwhelmed by a beta-adrenergic relaxing component. Administration of the beta-adrenergic receptor blocker, propranolol, during norepinephrine infusion caused similar increases in blood pressure in control, yohimbine-, and prazosin-treated rats. Uterine contractions, in contrast, were only significantly elevated during beta-adrenergic receptor blockade when yohimbine or imipramine had also been administered.     

Comparison of polymerase insertion and extension kinetics of a series of O2-alkyldeoxythymidine triphosphates and O4-methyldeoxythymidine triphosphate

The effect of alkyl group size on ability to act as deoxythymidine triphosphate (dTTP) has been studied for the carcinogen products O2-methyl-, O2-ethyl-, and O2-isopropyl-dTTP by using three types of nucleic acids as template and DNA polymerase I (Pol I) or Klenow fragment as the polymerizing enzymes. Apparent Km and relative Vmax values were determined in primer extension on M13 DNA at a single defined site, in poly[d(A-T)], and in nicked DNA. These data are the basis for calculation of the relative rate of insertion opposite A, relative to dTTP. The insertion rate for any O2-alkyl-dTTP is much higher than for a mismatch between unmodified dNTPs. Unexpectedly, O2-isopropyl-dTTP is more efficiently utilized than O2-methyl-dTTP or O2-ethyl-dTTP on each of the templates. O2-isopropyl-dTTP also substitutes for dTTP over extended times of DNA synthesis at a rate only slightly lower than that of dTTP. Parallel experiments using O4-methyl-dTTP under the same conditions show that it is incorporated opposite A more frequently than is O2-methyl-dTTP. Therefore, both the ring position and the size of the alkyl group influence polymerase recognition. Once formed, all O2-alkyl-T.A termini permit elongation, as does O4-methyl-T.A. In contrast to the relative difficulty of incorporating the O-alkyl-dTTPs, formation of the following normal base pair (C.G) occurs rapidly when dGTP is present. This indicates that a single O-alkyl-T.A pair does not confer significant structural distortion recognized by Pol I.     

The CYP2D gene subfamily: analysis of the molecular basis of the debrisoquine 4-hydroxylase deficiency in DA rats

The DA rat has been proposed as an animal model for the human debrisoquine 4-hydroxylase/bufuralol 1'-hydroxylase genetic deficiency. To determine the mechanism of this deficiency, we isolated and sequenced five cDNAs in the CYP2D gene subfamily including a new IID1 allele and two cDNAs of novel P450s, designated IID3 and IID5. IID3 and IID5 cDNA-deduced amino acid sequences contained 500 and 504 residues with calculated molecular weights of 56,683 and 57,081, respectively. IID5 displayed 20 amino acid differences with the IID1, yet bore only 72% and 76% similarity to IID2 and IID3. Despite an overall nucleotide similarity of 80-98% between the 4 cDNAs, a region of 134 nucleotides of sequence exists that contains only 1 base difference. This region is probably the result of gene conversion events between the P450 IID genes. Although all IID cDNAs were expressed into immunodetectable proteins using the COS cell SV40-based expression system, only IID1 could effectively catalyze the oxidation of the prototype substrate bufuralol. Expression of a cDNA isolated in an earlier study [Gonzalez, F. J., Matsunaga, T., Nagata, K., Meyer, U. A., Nebert, D. W., Pastewka, J., Kozak, C. A., Gillette, J., Gelboin, H. V., & Hardwick, J. P. (1987) DNA 6, 149-161], previously called db1 and now designated IID1v, produced a protein with a drastically reduced activity as compared to cDNA-expressed IID1 despite only four amino acid differences between the two cDNA-deduced protein sequences.(ABSTRACT TRUNCATED AT 250 WORDS)     

Transmitter release in hippocampal slices from rats with limbic seizures produced by systemic administration of kainic acid

The systemic injection of kainic acid (KA) has been shown to destroy neurons in the hippocampus and to induce limbic-type seizure activity. However, little is known on the neurochemical events that are associated with this convulsant effect. In the present work we studied the spontaneous and the K⁺-stimulated release of labeled -aminobutyric acid (GABA), glutamate, serotonin and dopamine, in hippocampal slices of KA-treated rats, at the moment of clinical seizures (2 h) and 72 h later. At the onset of convulsions we found a 40–45% decrease in the K⁺-stimulated release of GABA. The release of the other neurotransmitters was not significantly affected by KA treatment. After 72 h GABA release was still reduced by 30–40%. It is concluded that the epileptogenic effect of KA in the hippocampus is probably related to a diminished inhibitory GABAergic neurotransmission.     

Quantitative autoradiographic localization of neuropeptide Y (NPY) binding sites in rat posterior pituitary lobe

1. We have localized and quantified neuropeptide Y (NPY) binding sites in the rat pituitary gland after incubation of tissue sections in the presence of 125I-Bolton-Hunter NPY followed by autoradiography, computerized microdensitometry, and comparison to 125I-standards. 2. In the rat, NPY binding sites are localized exclusively to the part of the posterior pituitary lobe closer to the pituitary stalk. No NPY binding sites could be found in the intermediate or the anterior pituitary lobes. 3. Our results suggest a role for NPY in the regulation of pituitary function and, in particular, that of the neural lobe.