Postnatal development of mouse plasma and brain corticosterone levels: new findings contingent upon the use of a competitive protein-binding assay.

A competitive protein-binding (CPB) assay, suitable for measuring corticosterone levels in 20 mul of mouse plasma or 100 mg of brain, is described. The postnatal development of adrenocortical function was determined in C57BL/10 and DBA/1 mice by CPB assay of basal and stressinduced levels of plasma corticosterone and resting levels of brain corticosterone. Marked increases in both basal and stressed levels of plasma corticosterone were found beginning at day 12 after birth: mean basal levels rose from about 1 mug/u99 ml on day 12 to peak values of about 10-15 mug/100 ml on days 18-20, and then declined by day 30 to the 13-day level of 2.6 mug/100 ml. This pattern differs significantly from results obtained with standard fluorometric assays for corticosterone; it was determined that a major part of this discrepancy is due to the lack of specificity of the fluorometric assay. The developmental change in brain corticosterone was similar to the pattern found in plasma. Only the stress-induced levels of plasma corticosterone showed significant genetic variation, and this did not appear until about one week after the end of the relative stress-nonresponsive period. These findings should be useful in evaluating hypotheses concerning the developmental regulation of adrenocortical function and the action of glucocorticoids in regulating the biochemical differentiation of other tissues.     

Prospective assessment of positioning-related pain in robotic urologic surgery

This was a prospective study to assess positioning-related pain in 20 awake volunteers in the dorsal lithotomy (DL) and lateral decubitus (LD) positions. Each volunteer was put through the series of discrete, sequential steps used to achieve a final position; each step had two options. The Wong-Baker scale (WB) was used to rate pain for each option and the preferred option and ad lib comments were recorded. We found that awake volunteers could clearly and immediately distinguish differences in pain levels between position options. For the DL position, volunteers favored having the arms slightly flexed and pronated as opposed to being straight and supinated reflected by statistically less painful WB scores and option preference. Volunteers preferred having the neck flexed as opposed to being flat. For the LD position, volunteers reported statistically lower pain scores and preference for a foam roll for axilla support as opposed to a rolled blanket, the table flexed without the kidney rest as opposed to a raised kidney rest, and the over arm board as oppose to stacked blankets for contralateral arm support. Ad lib comments from the volunteers supported the above findings. To our knowledge, ours is the first study to demonstrate objective preferences for variations in surgical positioning using awake volunteers. This exercise with awake volunteers resulted in immediate changes in positioning for real robotic surgery patients in our practice.     

Neuropeptide-induced contraction and relaxation of the mouse anococcygeus muscle.

Isometric tension responses to neuropeptides were recorded from anococcygeus muscles isolated from male mice. This smooth muscle tissue is innervated by inhibitory nonadrenergic, noncholinergic nerves that resemble, ultrastructurally, the peptidergic neurons of the gastrointestinal tract; the physiological function of the anococcygeus is not known. Slow sustained contractions were produced by oxytocin (0.2-20 nM), [Arg8]vasopressin (0.4-200 nM), and [Arg]-vasotocin (0.4-100 nM); the mouse anococcygeus is, therefore, one of the few examples of nonvascular smooth muscle from male mammals to respond to low concentrations of oxytocin and related peptides. Substance P (0.5-8 microM) caused distinctive, biphasic increases in muscle tone of some, but not all, preparations. Other neuropeptides producing contractions were neurotensin (2-100 microM) and thyrotropin-releasing hormone (2-100 microM); the responses were of similar time course and displayed selective cross-desensitization, suggesting that these two peptides act through a common distinct mechanism. Tetradecapeptide somatostatin (10-80 microM) and its analog urotensin II (0.1-5 microM), a dodecapeptide from the urophysis of the teleost fish Gillichthys mirabilis, produced similar slowly developing relaxations of carbachol-induced tone. Piscine urotensin II, of which there are no reported effects on nonvascular mammalian systems, was 20-50 times more potent than somatostatin, a well-established mammalian hormone. Of the peptides studied, only vasoactive intestinal polypeptide (0.05-1 microM) caused rapid powerful relaxations in low concentrations; this is consistent with its proposed involvement in nonadrenergic, noncholinergic neurotransmission in the mouse anococcygeus.     

Total oxidant-scavenging capacities of plasma from glycogen storage disease type Ia patients as measured by cyclic voltammetry, FRAP and luminescence techniques

It has been suggested that the very low incidence of atherosclerosis in glycogen storage disease type Ia (GSD Ia) subjects might be attributed to elevated levels of uric acid, one of the potent low-molecular-weight antioxidants found in plasma. The present communication describes a use of two analytical methods—cyclic voltammetry and ferric reducing ability of plasma—and also two chemiluminescence methods to evaluate the total oxidant-scavenging capacities (TOSC) of plasma from GSD Ia patients. Our results verified the elevation of TOSC in GSD Ia patients and we propose the inclusion of luminescence and cyclic voltammetry assays as reliable methods for estimating TOSC in a variety of clinical disorders. Our findings with the cyclic voltammetry method add support to the assumption that the elevated uric acid levels might be the main contributor to plasma antioxidant capacity and possible protection against atherosclerosis.     

Plasminogen activator inhibitor-1 and vitronectin promote vascular thrombosis in mice

Occlusive thrombosis depends on the net balance between platelets, coagulation, and fibrinolytic factors. Epidemiologic information suggests that plasminogen activator inhibitor-1 (PAI-1), a central regulator of the fibrinolytic system, plays an important role in determining the overall risk for clinically significant vascular thrombosis. Vitronectin (VN), an abundant plasma and matrix glycoprotein, binds PAI-1 and stabilizes its active conformation. This study assessed the role of PAI-1 and VN expression in the formation of occlusive vascular thrombosis following arterial or venous injury. The common carotid arteries of 17 wild-type (WT) mice and 8 mice deficient in PAI-1 were injured photochemically while blood flow was continuously monitored. WT mice developed occlusive thrombi at 52.0 +/- 3.8 minutes (mean +/- SEM) following injury; mice deficient in PAI-1 developed occlusive thrombosis at 127 +/- 15 minutes (P <.0001). Mice deficient in VN (n = 12) developed vascular occlusion 77 +/- 11 minutes after injury, intermediate between the values observed for WT mice (P <.03) and mice deficient in PAI-1 (P <.01). PAI-1 and VN also affected the time to occlusion after injury to the jugular vein. Three WT mice developed occlusive venous thrombosis an average of 39.7 +/- 1 minutes following the onset of injury, whereas the jugular veins of 4 mice deficient in PAI-1 and 4 deficient in VN occluded 56.7 +/- 5 and 58.7 +/- 2 minutes, respectively, following injury (P <.04 and P <.01 compared to WT mice). These results suggest that endogenous fibrinolysis and its regulation by PAI-1 and VN have important roles in the development of occlusive vascular thrombosis after vascular injury. (Blood. 2000;95:577-580)     

Drug-induced thrombocytopenia is associated with increased binding of IgG to platelets both in vivo and in vitro

Thrombocytopenia is a common serious adverse effect of drug treatment. A variety of in vitro diagnostic techniques to confirm the diagnosis are available, but the majority lack sufficient sensitivity to detect all cases of drug-induced thrombocytopenia. We studied 19 patients with suspected drug-induced thrombocytopenia and demonstrated that platelet- associated IgG (PAIgG) was elevated in all at the time of thrombocytopenia, and PAIgG returned to normal levels as the thrombocytopenia resolved. In the majority of patients, the platelet count rapidly returned to normal after the drug was discontinued; however, in six patients, the thrombocytopenia persisted well beyond the period of time that the offending drug would be expected to be cleared from the blood. In 13 patients, serum obtained after recovery was used to identify the drug responsible for the thrombocytopenia in an in vitro assay. In all cases, the addition of the drug historically associated with the thrombocytopenic episode was associated with an increased binding of IgG to control platelets. For uncertain reasons, the concentration of drug required to increase the in vitro binding of IgG to test platelets was often more than the concentration usually achieved in vivo. Wider application of these techniques may provide better understanding of the clinical characteristics and mechanisms responsible for drug-induce thrombocytopenia.