The present study was designed to identify novel membrane proteins that signal during platelet aggregation. Because one putative mechanism for signaling by a membrane protein involves phosphorylation, we used oligonucleotide-based microarray analyses and mass spectrometric proteomics techniques to specifically discover membrane proteins and also identify those proteins that become phosphorylated on tyrosine, threonine, or serine residues upon platelet aggregation. Surprisingly, both techniques converged to identify a novel membrane protein we have termed PEAR1 (platelet endothelial aggregation receptor 1). Sequence analysis of PEAR1 predicts a type-1 membrane protein, 15 extracellular epidermal growth factor-like repeats, and multiple cytoplasmic tyrosines. Analysis of the tissue distribution of PEAR1 showed that it was most highly expressed in platelets and endothelial cells. Upon platelet aggregation induced by physiological agonists, PEAR1 became phosphorylated on tyrosine (Tyr-925), and serine (Ser-953 and Ser-1029) residues. PEAR1 tyrosine phosphorylation was blocked by eptifibatide, an ␣ IIb ␤ 3 antagonist, which inhibits platelet aggregation. Immune clustering of PEAR1 resulted in PEAR1 phosphorylation. Aggregation-induced PEAR1 tyrosine phosphorylation lead to the subsequent association with the ShcB adaptor protein. Platelet proximity induced by centrifugation also induced PEAR1 tyrosine phosphorylation, a reaction not inhibited by eptifibatide. These data suggest that PEAR1 is a novel platelet receptor that signals secondary to ␣ IIb ␤ 3 -mediated platelet-platelet contacts.Platelet aggregation during arterial thrombosis results in ischemic complications precipitating in acute myocardial infarction and stroke. Platelet aggregation is known to be mediated by signaling events initiated by primary platelet agonists such as thrombin, ADP, and collagen, which induce a conformational change in the platelet integrin ␣ IIb ␤ 3 , allowing it to bind soluble fibrinogen and von Willebrand factor, resulting in platelet cross-linking. Platelet-platelet contacts during aggregation subsequently initiate secondary signaling events. Aggregation-induced signaling can result in multiple platelet secondary signaling events such as calcium mobilization, protein tyrosine phosphorylations, cytoskeletal rearrangements, and the release of platelet-dense bodies and ␣-granules. Aggregation-induced signaling is key to the formation of stable aggregates, particularly when aggregation is induced by low concentrations of one or more primary agonists. Platelet activation also causes the release of ADP from dense bodies and the generation of thromboxane A2, both of which induce further platelet stimulation.Several mediators of aggregation-induced signals have been identified. One is ␣ IIb ␤ 3 itself, which becomes tyrosine-phosphorylated and also associates with numerous signaling and cytoskeletal proteins following platelet activation, allowing fibrinogen and/or von Willebrand factor binding and platelet aggregation. The importance o...

Sarcomere lengths were measured after pentobarbital anesthesia at five sites through the wall of the formaldehyde solution-fixed cadmium-arrested closed-chest rat left ventricle. Sections (250 micron) were cut from endocardium to epicardium with a freezing microtome. Selected sections were sonified, mixed with a gelatin-water solution, and placed on a glass slide. Sarcomere lengths were measured with an optical microscope at five sites through the wall. Sarcomere lengths progressively increased from section I (endocardium site) to section III (middle site) and IV. Sarcomere lengths were again shorter in section V (epicardium site). There was a progressive increase in sarcomere lengths with increasing intraventricular pressures. Sarcomere lengths did not significantly exceed optimum length.

Extracranial cerebrospinal fluid (CSF) shunt obstruction is one of the most important problems in hydrocephalus patient management. Despite ongoing research into better shunt design, robust and reliable detection of shunt malfunction remains elusive. The authors present a novel method of correlating degree of tissue ingrowth into ventricular CSF drainage catheters with internal electrical impedance. The impedance based sensor is able to continuously monitor shunt patency using intraluminal electrodes. Prototype obstruction sensors were fabricated for in-vitro analysis of cellular ingrowth into a shunt under static and dynamic flow conditions. Primary astrocyte cell lines and C6 glioma cells were allowed to proliferate up to 7 days within a shunt catheter and the impedance waveform was observed. During cell ingrowth a significant change in the peak-to-peak voltage signal as well as the root-mean-square voltage level was observed, allowing the impedance sensor to potentially anticipate shunt malfunction long before it affects fluid drainage. Finite element modeling was employed to demonstrate that the electrical signal used to monitor tissue ingrowth is contained inside the catheter lumen and does not endanger tissue surrounding the shunt. These results may herald the development of "next generation" shunt technology that allows prediction of malfunction before it affects patient outcome.

Shortening of the anterior papillary muscle of the left ventricle was demonstrated in six intact, tranquilized dogs. Two small metal markers that had been surgically implanted 3-50 months earlier were cineradiographically photographed during approximately ten sequential cardiac cycles in each of two orthogonal positions. Distances between markers were plotted for successive frames. The resulting curves were used to obtain maximum velocities of papillary muscle shortening and lengthening: 1.08 ± 0.29 muscle lengths/sec and 1.39 ± 0.48 muscle lengths/sec, respectively. From the two orthogonal planes, the average maximum spatial distance and the average minimum spatial distance between the markers were calculated. The mean percent shortening of 22.8 ± 6.5% was surprisingly large: it approximated the distance from the foot to the peak of the ascending limb of the myocardial lengthtension curve derived from isolated muscle studies. Mechanical studies on isolated papillary muscle have consistently shown reduced shortening with increasing loads. Since the in vivo dog papillary muscle has been reported to be under considerable tension during systole, there appears to be some contradiction between the degree of shortening found in the present study and the shortening observed in isolated papillary muscle studies. KEY WORDSmvocardial shortening length-tension curves cardiac metal markers myocardial velocities electrocardiogram• Papillary muscle has been investigated for two relatively independent reasons: (1) it is an experimentally convenient sample of ventricular myocardium, and (2) it is a clinically important portion of the functioning heart. Papillary muscle is easy to isolate and to remove from the ventricle. Its small diameter in some species and its participation during cardiac growth also contribute to its utility as an experimental material. Most papillary muscle studies have employed the isolated muscle (1-10), although more recently mechanical properties have been studied in the in situ muscle (11)(12)(13)(14). The essentially linear fiber arrangement within the muscle greatly simplifies the interpretation This work was supported in part by U. S. Public Health Service Grant 1 R01 HL-14651-01 from the National Heart and Lung Institute.A preliminary report on some of these results has been previously presented in abstract form (Fed Proc 27:2699, 1968.Received July 2, 1973. Accepted for publication September 6, 1974. Circulation Research, Vol. 36, January 1975 of data from both mechanical and microscopic studies (15).Papillary muscle function in the intact heart has, most often, been deduced indirectly from isolated muscle studies, clinical observations, autopsy reports, and theoretical considerations. Burch et al. (16) and Burch and Depasquale (17) have postulated that during ventricular systole the papillary muscle, by creating tension, acts as a stay to restrict mitral valve motion. When normal tension development does not occur, excessive mitral valve motion results in mitral regurgitation (the p...

Left ventricular dimensions were measured in Cd2+ arrested (presumably diastolic), open-chest rats. Aortic pressure was maintained at 137 cm H2O (100 mm Hg) and left-ventricular (luminal) pressures were established and maintained at their chosen values, each by means of reservoir systems. The selected left-ventricular pressures were chosen to be within or to even broaden the range of conceivable diastolic pressures (-3 to 48 cm H2O). After in situ fixation with 4% formaldehyde and gelatin embedding, the hearts were serially sectioned in the apex base direction to obtain information at 11 levels (10, 20, . . . 90, 100%). Tracings of selected sections were made along the edge of the left ventricular lumen and the pericardial surface. Volumes, surface areas, and mean external and internal radii of the left ventricle were derived. To quantify the circularity of sections a form factor (FF) was introduced (FF = 1 for a circular cross-section and less than one for other shapes). Ventricular lengths, radial dimensions, endocardial and epicardial surface areas, and total and luminal volumes increased with the increasing intraventricular pressures; as expected, the wall simultaneously thinned. Though its appearance was altered by the wall thinning, the curving muscle fascicular pattern was present over the entire pressure range examined. Endocardial surface areas increased more than did the epicardial surface areas. The endocardial FF value increased (more circular) at each section level as the pressure increased. The epicardial FF relationship was apparently constant (0.798 +/- 0.014) for all section levels from 10% through 90%, regardless of luminal pressure. These results, when taken in conjunction with the results of our previous published studies, prompted the following speculation. The wall of the diastolic ventricle is a fluid-filled chamber with intramyocardial pressures that may be higher than ventricular pressures.