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Biomedical subjects

K L Kirk

Publications and source records attributed to K L Kirk.

At least 127 records · Page 7Linked to original sources
PubMed1984

Quinacrine and basic amines in human platelets: subcellular compartmentation and effects on serotonin.

Human platelets appear to accumulate quinacrine both in a thrombin-releasable compartment (dense bodies or amine storage vesicles) and in another compartment from which it is released by agents known to collapse pH gradients (possibly lysosomes with an acidic interior). Approximately 61% of the total amount of quinacrine present in human platelets resides in dense bodies and 14+ in lysosomes, with the remainder probably present in the cytoplasm. Other basic amines are accumulated in the three compartments to widely varying extents, suggesting that several factors besides the existence of pH gradients act to determine the distribution of these substances within the cell. The fluorescence emission of quinacrine excited with 420 nm light is completely quenched for quinacrine inside both dense bodies and lysosomes, and the absorption of 440 nm light is decreased by approximately 25%. Quinacrine added to dense bodies at 37 degrees C induces the efflux of 5-hydroxytryptamine (5HT) from the bodies. There is, however, no 5HT loss following quinacrine entry at 0 degree C, and the relationship between the two types of amine movement varies according to incubation time at 0 degree C and 37 degrees C. This action of quinacrine therefore does not appear to be associated with stoichiometric exchange of 5HT and quinacrine, but rather to modulation of the passive permeability of the dense body membrane for 5HT.

Amines↗
PubMed1983

Synthesis and biological activity of 5-fluoroimidazole-TRH.

The 5-fluoroimidazole analogue of thyrotropin-releasing hormone, obtained by total synthesis from 5-fluoro-L-histidine, neither binds to rat pituitary cells nor stimulates release of prolactin from them. Lévine-Pinto et. al. reported an agonist, which was generated during presumptive photofluorination of the hormone and which they believed to be the 5-fluoro analogue. In addition to the striking contrast in biological activities, the chemical properties of the agonist differ markedly from those of our peptide and are inconsistent with expectation for the fluoroimidazole moiety. Despite its inactivity in pituitary functions, the authentic 5-fluoro analogue mimics the natural hormone with respect to cardiovascular responses in the central nervous system.

Animals↗
PubMed1983

Direct observation of 6-fluorodopamine in guinea pig nerve microsacs by 19F NMR.

19F nuclear magnetic resonance (NMR) has been used to examine the disposition of ring-fluorinated dopamine and norepinephrine in microsacs prepared from striata of guinea pig brains. Following incubation with a 10(-4)M initial concentration of 6-fluorodopamine (6F-DA), intact micmicrosacs at 4 degrees C gave a 19F NMR spectrum in which the 6F-DA present was sufficiently mobile to be visible. Intra-vesicular 6F-DA in striatal nerve terminals thus appears to exist in an environment resembling that in chromaffin vesicles but different from that prevailing inside the amine storage vesicles of platelets. Our data also suggest that the study of fluorinated compounds by 19F NMR can be used to expand our understanding of processes related to amine uptake, metabolism, and storage in nerves.

Animals↗
PubMed1983

Simultaneous determination of 3-methoxy-4-hydroxyphenylglycol, 5-hydroxyindoleacetic acid, and homovanillic acid in cerebrospinal fluid with high-performance liquid chromatography using electrochemical detection.

An improved high-performance liquid chromatographic method with electrochemical detection (HPLC-EC) for the simultaneous determination of 3-methoxy-4-hydroxyphenylglycol (MHPG), 5-hydroxyindoleacetic acid (5-HIAA), and homovanillic acid (HVA) in cerebrospinal fluid (CSF) of humans and nonhuman primates is described. Quantitation is based on the use of an internal standard, 5-fluoro-HVA. Sample preparation consists of mixing an aliquot of CSF with a solution of the internal standard followed by ultrafiltration. The precision of the method is high, with within-run and between-run coefficients of variation of 2-6% and less than 10%, respectively, in the concentration ranges of the metabolites encountered in human lumbar CSF. Accuracy was tested by comparing the present HPLC method with specific gas chromatographic-mass spectrometric (GS-MS) assays for MHPG and HVA and a GC-MS-validated HPLC assay for 5-HIAA: the correlations obtained were 0.968 for MHPG, 0.989 for 5-HIAA, and 0.999 for HVA, with no systematic bias between the methods. The use of ascorbate as a preserving agent for monoamine metabolites in CSF was not found to be necessary when proper care was exercised in sample handling and storage. The analysis of samples with up to 2% ascorbic acid was possible as well, but MHPG had to be assayed separately using an extraction procedure and an alternative internal standard, 3-ethoxy-4-hydroxyphenylglycol.

Animals↗
PubMed1983

Exploration of the role of phenolsulfotransferase in the disposition of serotonin in human platelets: implications for a novel therapeutic strategy against depression.

Human platelets have been used as models to evaluate a possible role for phenolsulfotransferase (PST) in the disposition of 5-hydroxytryptamine (5HT). The platelet M-type PST, which prefers biogenic amines as substrates, appears to be unique in its ability to O-sulfate dopamine and 4,6-difluoro-5HT (diF-5HT) at rates 6-20 times faster than 5HT. The assay technique employed can make a critical difference in the measured rates of sulfation of these amines, however. For example, use of purified alveolysin toxin to disrupt cell membranes can result in as much as a 20-fold increase in the observed velocity of 5HT O-sulfation. In intact platelets, only 1-3% of the total amount of radiolabelled material (as measured by a thin-layer chromatographic procedure) is the O-sulfate conjugate of 5HT. Up to 20% of the total H3-5HT taken up by cells becomes O-sulfated following treatments such as reserpine which lead to a large increase in the extra-vesicular (cytoplasmic) pool of H3-5HT. Availability of a compound producing irreversible inhibition of M-type PST activity would facilitate exploration of intra-platelet 5HT metabolism, but compounds reported to inhibit essentially 100% of the PST activity in other systems do not appear to interfere with the action of human platelet M-type PST. Because of the apparent similarity between PST present in human platelets and in nerve microsacs prepared from guinea pig brains, PST inhibitors effective in human platelets might also be valuable for studies of amine metabolism in brain tissue. Possible candidates are discussed. Furthermore, the behavioral effects of compounds proving to be PST inhibitors could provide novel therapeutic approaches to the problem of depressive illness.

Antidepressive Agents↗
PubMed1983

Basolateral potassium channel in turtle colon. Evidence for single-file ion flow.

Treatment of the apical surface of the isolated, ouabain-inhibited turtle colon with the polyene antibiotic amphotericin B permitted the properties of a barium-sensitive potassium conductance in the basolateral membrane to be discerned from the measurements of transepithelial fluxes and electrical currents. Simultaneous measurements of potassium currents and 42K fluxes showed that the movement of potassium was not in accord with simple diffusion. Two other cations, thallium and rubidium, were also permeable and, in addition, exhibited strong interactions with the potassium tracer fluxes. The results indicate that permeant cations exhibit positive coupling, which is consistent with a single-file mechanism of ion translocation through a membrane channel.

Animals↗
PubMed1983

Mechanism of epithelial lithium transport. Evidence for basolateral Na:Na and Na:Li exchange.

Measurement of transmural sodium fluxes across isolated, ouabain-inhibited turtle colon in the presence of a serosal-to-mucosal sodium gradient shows that in the absence of active transport the amiloride-sensitive cellular path contains at least two routes for the transmural movement of sodium and lithium, one a conductive path and the other a nonconductive, cation-exchange mechanism. The latter transport element can exchange lithium for sodium, and the countertransport of these two cations provides a mechanistic basis for the ability of tight epithelia to actively absorb lithium despite the low affinity of the basolateral Na/K-ATPase for this cation.

Animals↗
PubMed1983

Differential effects of fluoronorepinephrines on phosphatidylinositol turnover in rat pinealocytes.

(+/-)-Norepinephrines (NAs), substituted with fluorine at positions 2, 5, and 6 of the ring, were compared with the unsubstituted compound with respect to their capacity for eliciting increased incorporation of [32P]orthophosphate into phosphatidylinositol of pinealocytes. 5F-NA and 6F-NA were approximately equipotent with NA, whereas 2F-NA required a higher concentration and gave lower maximum stimulation. Inhibition of these effects by prazosin confirmed the participation of alpha 1-adrenergic receptors. The results are comparable with those reported for alpha 1-adrenergic receptor-mediated events in other systems and different from the beta-adrenergic receptor-mediated elevation of cyclic AMP in pinealocytes. These and earlier results emphasize the importance of the hydroxyl group at position 3 of the ring and at the beta-position of the side chain in catecholamine activation of the pineal alpha-adrenergic receptors, which are involved in alterations of phospholipid metabolism.

Animals↗
PubMed1983

6-Fluorocatecholamines as false adrenergic neurotransmitters.

In the present study 6-fluoronorepinephrine (6F-NE) is shown to be formed from 6-fluorodopamine (6F-DA) in vivo. The beta-hydroxylated fluorocompound is taken up by and stored in the adrenergic nerve terminals and can be released during sympathetic nerve stimulation. In the heart, the turnover rate of the exogenously administered 6F-NE was about the same as that of tritium-labeled norepinephrine. In the central nervous system, 6F-DA can be taken up by the nigrostriatal neurons. After depolarization of the dopaminergic neurons by potassium, 6F-DA is released along with the endogenous dopamine. Systemic administration of 6F-NE to the pithed rats produces dose-dependent increases in blood pressure but does not increase the heart rate. The vasopressor potency of 6F-NE is about the same as that of l-norepinephrine, about 2-fold greater than that of dl-norepinephrine. Combined treatment with yohimbine and prazosin antagonizes completely the vasopressor effect of 6F-NE. The duration of the pressor response to 6F-NE was twice that of dl- or l-norepinephrine. The present study indicates that 6-fluorocatecholamines fulfill the criteria for adrenergic false transmitters and may be useful in positron emission tomographic scanning for mapping specifically the adrenergic nervous system in the brain or in the peripheral sympathetic nerves.

Animals↗
PubMed1983

Microscopic methods for analysis of function in isolated renal tubules.

Using differential interference-contrast and fluorescence microscopy we have performed longitudinal observations of living, isolated and perfused nephron segments. With interference-contrast images we have made morphometric measurements (epithelial volume) and have compared those data to simultaneously measured physiologic parameters (delta psi T, Jv). Fluorescence images have allowed us to characterize qualitatively the distribution of a dye probe (acridine orange) within the epithelium and to follow changes in distribution in response to experimental maneuvers. In addition, preliminary results indicate that it should be possible to obtain quantitative information on the intracellular distribution of fluorescent probes, such as acridine orange, which have strong emission intensities. This should aid in the characterization of the cellular profiles of physiologic parameters such as pH or membrane potential. Collectively, these optical and physiological techniques should allow a meaningful analysis of structure and function in renal epithelia where a broad spectrum of transport processes can be examined.

Ammonium Chloride↗
PubMed1982

Syntheses and adrenergic agonist properties of ring-fluorinated isoproterenols.

2-Fluoro-, 5-fluoro-, and 6-fluoroisoproterenol were synthesized by reduction of the Schiff base formed between the corresponding ring-fluorinated 3,4-bis(benzyloxy)phenethanolamine and acetone, followed by reductive debenzylation in the presence of oxalic acid to yield crystalline neutral oxalates. The apparent beta-adrenergic potencies were determined in the isolated guinea pig atria. 2-Fluoro- and 5-fluoroisoproterenol were equipotent with (+/-)-isoproterenol, while 6-fluoroisoprotenol was virtually inactive. No alpha-adrenergic agonist activity (guinea pig aorta) was shown by any of the fluoroisoproterenols. Displacement of alpha- and beta-specific radioligands from isolated membrane preparations from rat brain by the fluoroisoproterenols were in agreement with the responses of the organ preparations. Thus, the apparent fluorine-induced specificity is due to specificity at the receptor binding site. The effects of fluorine substitution are discussed with regard to the apparent negative influence of the 6-fluoro substituent on the beta-agonist properties of isoproterenol, the lack of any increase in potency due to the 2-fluoro substituent, and the possibility of fluorine-induced changes in the electron density of the aromatic ring as a possible rational for the fluorine-induced specificity of both the fluoroisoproterenols and the fluoronoirepinephrines.

Adrenergic alpha-Agonists↗
PubMed1982

Uptake of 6-fluoro-5-hydroxytryptamine and 4,6-difluoro-5-hydroxytryptamine into releasable and non-releasable compartments of human platelets.

1 The ring-fluorinated compounds, 6-fluoro-5-hydroxytryptamine and 4,6-difluoro-5-hydroxytryptamine, are sterically similar to 5-hydroxytryptamine (5-HT) but have pKaS for their 5-hydroxyl groups (9.7 and 8.0 respectively) which are lower than that of 5-HT. 2 The rates at which [3H]-5-HT, [3H]-6-fluoro-5-HT, and [3H]-4,6-difluoro-5-HT entered the releasable (vesicular) and non-releasable compartments of washed human platelets during a 1 min incubation period at 37 degrees C were similar. These portions of the uptake process therefore appears to be relatively independent of the pKa of the 5-hydroxyl group. 3 The [3H]-4,6-difluoro-5-HT was unique since material accumulated in the non-releasable compartment during uptake did not migrate into the vesicular compartment under appropriate incubation conditions. 4 The data suggest that the zwitterionic form of 5-HT is not the transported species, but that the electronic configuration about the 5-hydroxyl group may be critical for translocation from a non-releasable to a releasable compartment.

Blood Platelets↗
PubMed1982

Noradrenergic responses in rat hippocampus: electrophysiological actions of direct- and indirect-acting sympathomimetics in the in vitro slice.

The effects of perfusion of postulated direct- and indirect-acting sympathomimetics on evoked potentials in the CA1 region of the in vitro rat hippocampus were examined. A selective alpha agonist, 6-fluoronorepinephrine, produced depressions of population spike amplitude which were antagonized by the alpha antagonist phentolamine, but not by the beta antagonist timolol. The selective beta agonist, 2-fluoronorepinephrine, produced increases in population spike amplitude which were antagonized by timolol but not by phentolamine. Weak and variable responses were seen to the indirect-acting sympathomimetic tyramine, with lower doses producing increases and higher doses producing decreases in population spike amplitude, respectively. As with 2-fluoronorepinephrine, increases in spike amplitude elicited by tyramine were blocked by timolol but not by phentolamine. Another indirect-acting sympathomimetic, d-amphetamine, produced only increases in population spike amplitude which were blocked by timolol. Phencyclidine, an agent which may produce some of its central effects via noradrenergic synapses, was virtually ineffective in producing catecholamine-like responses in this system. Only nonspecific, local anesthetic effects were observed. Taken together with previous studies, these results support the hypothesis that activation of alpha and beta receptors decreases and increases, respectively, pyramidal cell excitability. Furthermore, although both alpha and beta receptors appear to be capable of interacting with endogenously released norepinephrine, the beta response may predominate.

Action Potentials↗
PubMed1981

Influence of fluorine substitution on the site of enzymatic O-methylation of fluorinated norepinephrines.

The extent of meta- and para-O-methylation by catechol O-methyltransferase of 2-fluoro-, 5-fluoro-, and 6-fluoronorepinephrine (FNE) at pH 7 and 9 was determined. The rank order of preference for para-O-methylation is 5FNE much greater than NE greater than 6FNE greater than 2FNE. In all cases, increasing the pH to 9 results in an increase in para-O-methylation. Results with 2F- and 5FNE demonstrate the importance of ionization in the methyltransferase reaction when fluorine is situated ortho to one of the phenolic groups. To establish unequivocally the identities of the products, the isomeric, monofluorinated vanillins and isovanillins were synthesized and directly related to the products formed enzymatically from the monofluorinated norepinephrines.

Catechol O-Methyltransferase↗
PubMed1981

Anomalous compartmentation of 5-hydroxytryptamine in intact human platelets.

1 The movement of radiolabelled 5-hydroxytryptamine (5-HT) between the extracellular medium, the thrombin-releasable (vesicular) compartment, and a non-thrombin-releasable compartment has been investigated in washed human platelets. 2 Appreciable amounts of extracellular 5-HT can accumulate in a non-releasable compartment. Depending on the incubation conditions and the amount of 5-HT already present inside the cell, non-releasable 5-HT can either remain non-releasable or rapidly migrate into the vesicular compartment. 3 Measurable amounts of vesicular 5-HT can also enter a non-releasable compartment. However, vesicular 5-HT which becomes non-releasable does not appear to mix with 5-HT becoming non-releasable following uptake from the extracellular medium. 4 Extracellular 5-HT can be added to vesicles and to the non-releasable compartment in the presence of appreciable quantities of 5-HT already in one or both compartments. 5 The vesicular and non-releasable compartments appear to accumulate 5-HT independently of one another. Furthermore, most vesicular accumulation of 5-HT occurs from the extracellular medium, rather than by translocation of non-releasable 5-HT into the vesicular compartment.

Biological Transport↗