PET neuroimaging of clomipramine challenge in humans: focus on the thalamus.

PET neuroimaging of serotonin responsivity relied previously mainly on fenfluramine, but that drug has been withdrawn from the market. Therefore, we determined whether clomipramine, which stimulates serotonergic mechanisms by inhibiting serotonin reuptake, has reliable effects in the healthy human brain as measured by [15O]H2O PET. The clomipramine challenge markedly reduced the relative rate of blood flow in the selected region of interest, namely the dorsomedial nucleus of the thalamus, a limbic region rich in serotonin uptake sites. These findings show similarities between effects of fenfluramine and clomipramine in the healthy human brain, and support the use of the clomipramine challenge in conjunction with PET for studying cerebral serotonergic mechanisms.

Neuroimaging of serotonin uptake sites and antidepressant binding sites in the thalamus of humans and 'higher' animals.

This review presents the results of in vitro, ex vivo and in vivo studies carried out primarily for identifying serotonin uptake sites and/or antidepressant binding sites in the brain of humans and 'higher' animals, namely nonhuman primates and pigs. Five lines of evidence are considered. First, studies carried out in vitro using synaptosomes or membrane preparations from human, nonhuman primate, and porcine brain have shown that certain thalamic nuclei are major sites of serotonergic neurotransmission in these species. Second, studies carried out in vitro or ex vivo using autoradiography or immunohistochemistry have indicated that the dorsomedial nucleus and some adjacent regions of the thalamus have a particularly high density of binding sites for antidepressant drugs that are selective serotonin reuptake inhibitors (SSRIs). Third, studies carried out in the living brain of nonhuman primates and pigs have found that SSRIs, radiolabeled for use in PET or SPECT, accumulate to a relatively high degree in midline and dorsal nuclei of the thalamus. Fourth, studies carried out using PET or SPECT radioligands in humans have demonstrated that regions in and around the dorsomedial nucleus of the thalamus are principal sites for accumulation of SSRIs. Fifth, studies of behavior of humans suffering from localized tissue damage in the thalamus have reported that symptoms of mania often occur in the patients, in accordance with the notion that an intact thalamus is required for normal regulation of mood. Taken together, the findings are consistent with the hypothesis that serotonergic neurotransmission in the dorsomedial nucleus of the thalamus could be causally involved in the pathophysiology of affective disease as well as in therapeutic actions of SSRIs.

Maternal iodine status and thyroid volume during pregnancy: correlation with neonatal iodine intake.

Differences in pregnancy-associated alterations in thyroid volume and urinary iodine (UI) excretion have been attributed to geographical variations in dietary iodine intake. In this study, ultrasound-measured thyroid volume and UI excretion were assessed during the 3 trimesters of pregnancy, at delivery, and at 6 weeks postpartum. Urine specimens also were obtained from mothers and both breast- and formula-feeding infants at 3 days after delivery. Thyroid volume showed a significant increase (maximum 47.0%), compared with nonpregnant control values over the 3 trimesters of pregnancy, which occurred as early as the first trimester and was paralleled by increased UI excretion, followed in turn by a precipitous fall at delivery. UI excretion in breast-feeding neonates (100 +/- 6.8 micrograms/L) was significantly higher than in their mothers (76 +/- 5.6 micrograms/L; p < 0.01) but was significantly lower (43 +/- 3.5 micrograms/L) in formula-fed infants. The results suggest that in an area of moderate dietary iodine intake, UI loss during pregnancy may result in maternal thyroid enlargement. The ability of the breast to transport iodine compensates for this loss in breast-fed infants, but this protection may be lost in formula feeding.

A pathway of multi-chaperone interactions common to diverse regulatory proteins: estrogen receptor, Fes tyrosine kinase, heat shock transcription factor Hsf1, and the aryl hydrocarbon receptor.

A variety of regulatory proteins, including different classes of transcription factors and protein kinases, have been identified in complexes with Hsp90. On careful examination of unactivated progesterone receptor complexes, eight different protein participants have been identified, and each can be considered a component of the cytoplasmic molecular chaperone machinery. These proteins are Hsp90, Hsp70, Hip, p60, p23, FKBP51, FKBP52 and Cyp40. Studies in a cell-free assembly system have helped to define a highly ordered, dynamic pathway for assembly of progesterone receptor complexes. In the present study, target proteins other than progesterone receptor were used in this cell-free system to assemble complexes in vitro and to compare the composition of resulting complexes. Targets used were human estrogen receptor, human Fes protein-tyrosine kinase, human heat shock transcription factor Hsf1, and human aryl hydrocarbon receptor. The striking similarity of resulting target complexes with previously characterized progesterone receptor complexes suggest that each of these targets undergoes a common assembly pathway involving multiple chaperone components in addition to Hsp90.

Cost-benefit of combined use of acupuncture, Shiatsu and lifestyle adjustment for treatment of patients with severe angina pectoris.

Sixty-nine patients with severe angina pectoris were treated with acupuncture, Shiatsu and lifestyle adjustments, and were followed for 2 years. Forty-nine patients were candidates for coronary-artery bypass grafting (CABG), whereas bypass grafting was rejected in the remaining 20 patients. We compared our endpoint findings with those of a large prospective, randomized trial comparing CABG with percutaneous transluminal coronary angioplasty (PTCA). The incidence of death and myocardial infarction was 21% among the patients undergoing CABG, 15% among the patients undergoing PTCA and 7% among our patients. No significant difference was found concerning pain relief between the three groups. Invasive treatment was postponed in 61% of our patients due to clinical improvement, and the annual number of in-hospital days was reduced by 90%, bringing about an estimated economic saving of 12,000 US $ for each of our patients. Despite the fact that the men in the present study, had significantly less positive expectations towards the outcome of the treatment, when compared to the women, there was no significant difference concerning the effect. The study suggests that the combined treatment with acupuncture, Shiatsu and lifestyle adjustment may be highly cost effective for patients with advanced angina products.

Effect of flunarizine and calcium on serotonin uptake in human and rat blood platelets and rat synaptosomes.

Calcium and the calcium overload blocker flunarizine exert profound effects on mood. We therefore studied the effect of calcium and flunarizine on serotonin uptake in human and rat blood platelets and in rat synaptosomes. Calcium (1.3 mmol/L) had a weak inhibiting effect on serotonin uptake in blood platelets, whereas no effect was observed in synaptosomes. Flunarizine inhibited serotonin uptake in a concentration dependent manner with an IC50 value of 1 mumol/L in blood platelets and 5 mumol/L in synaptosomes. The inhibition did not depend on the presence of extracellular calcium indicating that the effect is not coupled to a blockade of cellular calcium influx. In human blood platelets, the inhibition was of the noncompetitive type. These results indicate that flunarizine interacts directly with the 5-HT uptake site. The relatively high concentration of flunarizine required to inhibit 5-HT uptake may question the clinical importance of this effect.

Abomasal infusion of casein enhances nitrogen retention in somatotropin-treated steers.

The effects of bovine somatotropin (bST) administration and abomasal protein infusion on nitrogen metabolism were investigated using four abomasally-cannulated Holstein steers in a 4 x 4 Latin square design. Treatments were factorially arranged to include daily abomasal infusion (water or casein, 0.9 g/kg body wt) and daily bST injection (0 or 200 micrograms/kg body wt). Each treatment period lasted for 23 d and consisted of an adjustment period (d 1 to 7), casein infusion (d 8 to 23) and bST treatment (d 10 to 23). Nitrogen metabolism and serum insulin-like growth factor-I (IGF-I) profiles were characterized on d 16-22. Animals were fed a total mixed diet balanced to achieve 0.75 kg gain/d and supplemented to provide 115% of requirements for crude protein and minerals. Nitrogen retention was increased by casein and bST individually (21 and 33%, respectively) or in combination (75%). Somatotropin treatment increased the biological value of absorbed N, which was consistent with reductions in plasma urea nitrogen. Circulating IGF-I was increased by bST and was highly correlated (r = 0.73) with N retention. Overall, bST altered requirements so that less absorbed N was required per unit of N retained. However, results also demonstrated that because of limitations in amino acids supplied from microbial and ruminal escape protein in young growing cattle, amino acid supply may limit the magnitude of N retention response to bST under many situations.

Use of recombinant biotinylated aequorin in microtiter and membrane-based assays: purification of recombinant apoaequorin from Escherichia coli.

Aequorin is a calcium-dependent bioluminescent protein isolated from the hydromedusan Aequorea victoria. The gene for aequorin has been cloned and overexpressed in Escherichia coli [Prasher et al. (1985) Biochem. Biophys. Res. Commun. 126, 1259; Prasher et al. (1987) Biochemistry 26, 1326]. Higher levels of expression have recently been obtained by subcloning aequorin cDNA into the pRC23 plasmid vector such that its expression is under control of the lambda PL promoter [Cormier et al. (1989) Photochem. Photobiol. 49, 509]. Purification of recombinant apoaequorin from E. coli containing this new recombinant plasmid (pAEQ1.3) was accomplished by a two-step procedure involving gel filtration and anion-exchange chromatography on Sephadex G-100 and DEAE-Sepharose, respectively. Typically, 400-500 mg of recombinant protein was obtained from 100 L of fermentation culture. The purified recombinant apoaequorin could be converted to aequorin in high yield upon incubation with synthetic coelenterate luciferin, dissolved oxygen, and a thiol reagent with a photon yield similar to the native photoprotein. Detection of recombinant aequorin in the Dynatech ML1000 Microplate luminometer was linear between 10(-18) and 10(-12) mol, and little loss of specific activity was observed when the protein was derivatized with biotin. The biotinylated derivative was stable when frozen, lyophilized, or stored at 4 degrees C. The feasibility of using biotinylated aequorin as a nonradioactive tag was established by its application in a variety of solid-phase assay formats using the high-affinity streptavidin/biotin interaction. A microtiter-based bioluminescent immunoassay (BLIA) using biotinylated aequorin and the ML1000 luminometer was developed for the detection of subnanogram amounts of a glycosphingolipid (Forsmann antigen). In addition, nanogram to subnanogram quantities of protein antigens and DNA, immobilized on Western and Southern blots, respectively, were detected on instant and X-ray films using biotinylated aequorin.

Assembly of progesterone receptor with heat shock proteins and receptor activation are ATP mediated events.

To better understand assembly mechanisms of progesterone receptor (PR) complexes, we have developed a cell-free system for studying PR interactions with the 90- and 70-kDa heat shock proteins (hsp90 and hsp70), and we have used this system to examine requirements for hsp90 binding to PR. Purified chick PR, free of hsp90 and immobilized on an antibody affinity resin, will rebind hsp90 in rabbit reticulocyte lysate when several conditions are met. These include: 1) absence of progesterone, 2) elevated temperature (30 degrees C), 3) presence of ATP, and 4) presence of Mg2+. We have obtained maximal hsp90 binding to receptor when lysate is supplemented with 3 mM MgCl2 and an ATP-regenerating system. ATP depletion of lysate by dialysis or by enzymatic means blocks hsp90 binding to PR; likewise, addition of EDTA to lysate blocks hsp90 binding, but binding is restored by the addition of excess Mg2+. Addition to lysate of monoclonal antibody against hsp70 inhibits hsp90 binding to PR and destabilizes preformed complexes. Stabilization of hsp90-receptor complexes also requires ATP, indicating that ATP and hsp70 are needed to form and to maintain hsp90 complexes. Hormone-dependent activation of reconstituted receptor complexes was also examined. The addition of progesterone to the reticulocyte lysate promotes dissociation of hsp90 and hsp70 from the receptor. This also appears to require ATP and dissociation is most efficient in the presence of an ATP-regenerating system. In conclusion, these studies indicate that PR-hsp90 complexes do not self-assemble; instead, assembly is probably a multistep process requiring ATP and other cellular factors.

Micronucleated erythrocytes as an index of cytogenetic damage in humans: demographic and dietary factors associated with micronucleated erythrocytes in splenectomized subjects.

Erythrocytes containing micronuclei serve as an indicator of genotoxic exposure in splenectomized individuals. Micronucleated erythrocytes, derived from cytogenetically damaged RBC precursors, are not selectively removed from peripheral blood in individuals who lack splenic function. The relationship between micronucleated cell frequencies and demographic, environmental, and dietary factors was examined in 44 subjects with previous splenectomy due to trauma. Their micronucleated cell counts fit a log-normal distribution, with geometric means of 3.3 micronucleus-containing cells/1000 reticulocytes and 2.7/1000 normochromatic erythrocytes. A multiple regression analysis showed that drinking five cups of coffee or tea/day (relative to none) was associated with an approximately 2-fold higher frequency of micronucleated cells. Weaker statistical associations were also noted with micronucleus frequency and the consumption of calcium supplements (associated with a higher frequency) and vitamins A, C, or E (lower frequency). An apparent trend of higher micronucleus counts with age was attenuated when other factors were considered in the regression. Cigarette smoking and decaffeinated coffee consumption were among the factors not associated with elevated micronucleated cell frequencies. Because the occurrence of micronuclei in reticulocytes reflects cytotoxic exposures within the past 3-8 days, it may be possible to test directly the relationship of these factors to micronucleus formation through intervention studies.

Contrast enhancement technique for dynamic hepatic computed tomographic scanning.

A study comparing precontrast and survey postcontrast dynamic computed tomographic (CT) scanning was performed on 60 patients who had suspected hepatic metastases. An incremental dynamic technique was used during and following a 50-g iodine load administered over two minutes. The survey postcontrast dynamic technique was superior in both sensitivity and contrast differentiation and yielded no known false-negative examinations. No postprocedure renal dysfunction was observed. High-dose contrast-material delivery in conjunction with incremental dynamic CT scanning appears to be the most suitable technique for performing postcontrast hepatic CT examinations.

Lithium and rubidium: effects on the rhythmic swimming movement of jellyfish (Aurelia aurita).

The effects of adding LiCl, RbCl, KCl or NaCl to sea water at concentrations up to 30 mmoles/1 on the frequency of contraction of jellyfish (Aurelia aurita) suggest that studies on phylogenetically low animals with relatively simple nervous systems may be of use to determine mechanisms of action of lithium and rudidium on movements.