Alternative splice isoforms of small conductance calcium-activated SK2 channels differ in molecular interactions and surface levels

Small conductance Ca²⁺-sensitive potassium (SK2) channels are voltage-independent, Ca²⁺-activated ion channels that conduct potassium cations and thereby modulate the intrinsic excitability and synaptic transmission of neurons and sensory hair cells. In the cochlea, SK2 channels are functionally coupled to the highly Ca²⁺ permeant α9/10-nicotinic acetylcholine receptors (nAChRs) at olivocochlear postsynaptic sites. SK2 activation leads to outer hair cell hyperpolarization and frequency-selective suppression of afferent sound transmission. These inhibitory responses are essential for normal regulation of sound sensitivity, frequency selectivity, and suppression of background noise. However, little is known about the molecular interactions of these key functional channels. Here we show that SK2 channels co-precipitate with α9/10-nAChRs and with the actin-binding protein α-actinin-1. SK2 alternative splicing, resulting in a 3 amino acid insertion in the intracellular 3′ terminus, modulates these interactions. Further, relative abundance of the SK2 splice variants changes during developmental stages of synapse maturation in both the avian cochlea and the mammalian forebrain. Using heterologous cell expression to separately study the 2 distinct isoforms, we show that the variants differ in protein interactions and surface expression levels, and that Ca²⁺ and Ca²⁺-bound calmodulin differentially regulate their protein interactions. Our findings suggest that the SK2 isoforms may be distinctly modulated by activity-induced Ca²⁺ influx. Alternative splicing of SK2 may serve as a novel mechanism to differentially regulate the maturation and function of olivocochlear and neuronal synapses.     

Injury to the glenoid fossa

An illustrated case of fracture through the confines of the glenoid fossa by the mandibular condyle is presented. The mechanism of injury and postinjury sequelae of this rarely reported but frequently suspected occurrence are discussed and a treatment rationale proposed. The key to the diagnosis of this entity is an awareness of vector forces directed to the region of the glenoid fossa with or without evidence of subcondylar fracture.     

Beiträge zur Embryonalentwicklung vonLimulus Polyphemus L. (Chelicerata,Xiphosura)

Zusammenfassung Es wurde die Entwicklungsgeschichte einiger Organsysteme vom Beginn der Segmentierung des Keimstreifens bis zum 1. Larvenstadium anLimulus polyphemus untersucht.Von besonderem Interesse erschien die Ausgestaltung metamerer Strukturen mesodermaler und ektodermaler Herkunft.Zwei Fragen standen im Vordergrund dieser Arbeit, einmal die umstrittene Abgrenzung der Prosoma-Opisthosoma-Grenze, zum zweiten die Entstehung des Syncephalon eines ursprünglichen Cheliceraten. Die Mesodermversorgung des Kopfes entspricht der anderer genauer untersuchter Cheliceraten, Es ist labrales Mesoderm und ein ausgedehntes prächelicerales Coelom vorhanden. Die prospektive Bedeutung des labralen Mesoderms ist die Ausbildung der schwachentwickelten eigentlichen Labralmuskulatur, vor allem aber die Bildung von Mundwinkelprotraktoren, rostralen Vorderdarmdilatatoren und Vorderdarmmuskulatur.Die prospektive Bedeutung des Prächelicerencoeloms liegt in der Bildung der Aorta anterior. Vermutlich ist die Entstehung des vorderen komplexen Dotterseptums ebenfalls auf den Einfluß dieses Prächelicerencoeloms zurückzuführen.Im Verlauf dieses Dotterseptums erfolgt eine bemerkenswerte Umkehrung der serial homolog entstehenden Anlagen mesodermaler Elemente. Die dorsale Extremitätenmuskulatur der Cheliceren gelangt gegenüber der dorsalen Extremitätenmuskulatur des folgenden, ja sogar des übernächsten Segmentes in eine caudal verschobene Position. Wahrscheinlich ist vergleichbares auch für die vordersten Suspensormuskeln des Endosternits der Fall.Im Nervensystem dieses Bereiches lassen sich ebenfalls eine Fülle von Besonderheiten nachweisen:Das Ganglion der Cheliceren besitzt nur eine vor dem Vorderdarm verlaufende Kommissur. Unabhängig von ihm entsteht in der Vorderdarmseitenwand ein Strang neurogenen Gewebes, der sich sekundär mit dem Chelicerenganglion verbindet und die sogenannten Rostralganglien, besser Stomodaealganglien oder Pharyngealganglien liefert, von denen aus, der Seitenwand des Vorderdarmes anliegend, sehr deutliche Nervenstränge dem Vorderdarm entlang ziehen (Stomodaealnerven nach Patten u. Redenbaugh, 1900).Einmalig innerhalb der Arthropoden ist die Versorgung des Vorderdarmes mit Muskulatur, die Beziehung zu sicher poststomodaeal angelegten Segmenten aufweist. Wenn auch deren Herkunft nur in wenigen Fällen geklärt werden konnte, so ist zumindest sicher, daß poststomodaeale Anteile des Primärsternits in den Vorderdarm eingebaut werden.Vor dem Chelicerenganglion liegt das von Johansson (1933) als Antennalganglion beschriebene Gebilde. Seine Bedeutung wird diskutiert. Rostral folgt ein weiteres Ganglion, welches aus den sekundär verschmolzenen Zentralkörperanlagen entsteht. Das Archicerebrum besteht aus den sehr spät entstehenden und mit zipfelförmigen Fortsätzen bis ins Epistom reichende Corpora pedunculata, dem sogenannten Cerebralganglion, welches die Sehzentren für die Medianaugen enthält und den seitlich angrenzenden optischen Ganglien. Das Gehirn vonLimulus ist durch Konzentrationsprozesse in medianer Richtung und damit verbunden einer Emporhebung von Zentralkörperganglion und Antennalganglion gekennzeichnet.Für das Verständnis der ungewöhnlichen Position des Komplexauges vonLimulus ist der Nachweis seiner caudalen Verlagerung wichtig.Das Herz entsteht aus den dorsalen Teilen der Coelome 5 bis 13.Das 7. Metamer wird ganz und das 8. zum größten Teil in das Prosoma einbezogen. Eine entscheidende Bedeutung der sich hierbei abspielenden Verlagerungsvorgänge kommt dem Dottersegment 6 zu. Im Opisthosoma verbleiben die Ganglien 9 bis 16. Die Ausgestaltung dorsoventraler Muskulatur macht die Anlage von insgesamt 18 Metameren wahrscheinlich. Die Zuordnung des Coeloms zu ektodermalen Strukturen (Seitenzähne und Seitenstachel des Opisthosomas sowie dorsalen Borsten wird diskutiert).Die anLimulus beobachteten gegenläufigen Gestaltungsbewegungen stellen eine Fülle von Fragen hinsichtlich der sie bewirkenden Faktoren.

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Contributions to the embryology ofLimulus polyphemus L. (Chelicerata, Xiphosura)

Summary The embryology of some organ systems from the beginning of the segmentation of the germ band to the first larval stage has been studied onLimulus polyphemus.The main concern was the formation of the metameric structures of mesodermal and ectodermal origin.Two question were mainly dealt with in this work: firstly, the controversal limitation between prosoma and opisthosoma; secondly, the development of the syncephalon of a primitive chelicerate. The mesodermal supply of the head agrees with other carefully studied chelicerates. There is a labral mesoderm and an expanded precheliceral coelom. The prospective importance of the labral mesoderm is the formation of the weakly developed labral muscles s.str., but mainly the development of the protractors of the mouth edge, the rostral dilatators of the esophagus, and the muscular sheath of the esophagus.The prospective significance of the precheliceral coelom is the formation of the aorta anterior. Presumably the development of the most anterior compound yolk septum is also due to the influence of this precheliceral coelom.In the course of this yolk septum a remarkable distortion of serially homologous developing anlagen of mesodermal elements occurs. The dorsal muscular system of the chelicera is placed caudal to the first ambulatory limb, even caudal to that of the second ambulatory limb. It might be the same for the anterior suspensor muscles of the endosternite.In the nervous system of this region quite a number of peculiarities can be found: The ganglion of the cheliceres has only one commissure which is in front of the esophagus. Independent of the cheliceral ganglion a line of neurogenic tissue develops in the lateral walls of the esophagus. Later this line comes into connection with the pilem of the cheliceral ganglion. This neurogenic line forms the so-called rostral, or better stomodaeal, ganglion and the very distinct stomodaeal nerves.Unique within the arthropods is the supply of the esophagus with dilatator muscles, which go to the base of the ambulatory legs 1 and 4 and the endosternite in close connection with the endosternocoxal muscles of those legs. The origin of most of the muscles is not quite clear but some show that poststomodaeal parts are incorporated in the esophagus.In front of the cheliceral ganglion we find the antennal ganglion first described by Johansson (1933). Its importance is discussed. An other ganglion follows further rostral which develops out of the secondarily united anlagen of the central body. The archicerebrum contains the corpora pedunculata which extends backward into the epistome, the so-called cerebral ganglion with the centers for the dorsomedian eyes, and laterally the optic ganglion. The brain ofLimulus is emphasized by the process of concentration in a median direction which brings the central body and the antennal ganglion in dorsal position.For an understanding of the unusual position of the complex eye ofLimulus, the demonstration of its caudal shifting is of importance.The heart develops out of dorsal parts of coelomic cavities 5 to 13.The seventh segment is completely amd the eighth segment in its main parts incorporated in the prosoma. The enlargement of the sixth yolk segment plays a dominant rôle within the process of shifting. Ganglions 9 to 16 stay within the opisthosoma. The development of the dorsoventral muscles shows the formation of 18 metamers. The relation between the coelomic and extodermal structures such as the spine, teeth, and apodemes of the opisthosoma is discussed.The shifting of material in a different or often antagonistic direction raises many question concerning the physiologic factors.

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Verzeichnis der Abkürzungen Aa Aorta anterior - AG Antennalganglion - ALat Arteria lateralis - Btm Branchiothorakalmuskeln,=Branchioprosomamuskeln - Ce Cerebrum - CG Cerebralganglion - Ch Chelicere - ChG Chelicerenganglion - Chi Chilarium - ChN Chelicerennerv - Cö Coelom - CöP Coelom der Prächelicere - Cp Corpora pedunculata - Cpa Caudalpapille - Cx Coxa - d dorsal - Dd Dotterdivertikel - Dlm Dorsaler Längsmuskel - Dprm Dorsaler Promotormuskel - Drm Dorsaler Remotormuskel - Ds Dottersegment - Dvm Dorsoventralmuskel - E Endosternit - Ent Entapophyse - Fe Femur - Fl Flabellum - G Ganglion - H Herz - Hv Herzventil - h hinterer - i innerer - KA Komplexauge - Ki Kiemenblätter - Kkn Kiemenknorpel - Ko Kommissur - KoCh Kommissur der Chelicere - Kv Kiemenvene - l lateraler - La Lade - Lam Longitudinaler abdominaler Muskel - Ln Labralnerv - Lo Lateralorgan - MA Medianauge - MAA Medianaugenanlage - MAE Medianaugeneinstülpung - MASt Medianaugenstrang - Mes Mesoderm - MuCh dorsale Chelicerenmuskeln - N Nerv - NCh Chelicerennerv - NMA Medianaugennerv - NZ Neurale Zwischenzellen - OG Optisches Ganglion - Ol Oberlippe - Olmes Oberlippenmesoderm - P Pedunculus - Pi Pilem - Pt Patella - Py pyknotische Kerne - SA Seitenarterienanlage - SuE Suspensormuskeln des Endosternits - Sch Schildrand - SchA Schildrandanlage - Scl Sclerite der Coxa - Sto Stomodaeum - Stomes Stomodaeummesoderm - StoG Stomodaeumganglion - StZ Stäbchenzellen - Ta Tarsus - Tb Tibia - Tr Trochanter - v ventrale - Vd Vorderdarm - VLm ventrale Längsmuskel - Vo Ventralorganartige Bildung - Vpkm Venoperikardialmuskel - Zk Zentralkörperganglion     

Modeling interactions between voltage-gated Ca2+ channels and KCa1.1 channels

High voltage-activated (HVA) Cav channels form complexes with KCa1.1 channels, allowing reliable activation of KCa1.1 current through a nanodomain interaction. We recently found that low voltage-activated Cav3 calcium channels also create KCa1.1-Cav3 complexes. While coimmunoprecipitation studies again supported a nanodomain interaction, the sensitivity to calcium chelating agents was instead consistent with a microdomain interaction. A computational model of the KCa1.1-Cav3 complex suggested that multiple Cav3 channels were necessary to activate KCa1.1 channels, potentially causing the KCa1.1-Cav3 complex to be more susceptible to calcium chelators. Here, we expanded the model and compared it to a KCa1.1-Cav2.2 model to examine the role of Cav channel conductance and kinetics on KCa1.1 activation. As found for direct recordings, the voltage-dependent and kinetic properties of Cav3 channels were reflected in the activation of KCa1.1 current, including transient activation from lower voltages than other KCa1.1-Cav complexes. Substantial activation of KCa1.1 channels required the concerted activity of several Cav3.2 channels. Combined with the effect of EGTA, these results suggest that the Ca²⁺ domains of several KCa1.1-Cav3 complexes need to cooperate to generate sufficient [Ca²⁺]_i, despite the physical association between KCa1.1 and Cav3 channels. By comparison, Cav2.2 channels were twice as effective at activating KCa1.1 channels and a single KCa1.1-Cav2.2 complex would be self-sufficient. However, even though Cav3 channels generate small, transient currents, the regulation of KCa1.1 activity by Cav3 channels is possible if multiple complexes cooperate through microdomain interactions.     

Genetic variation in the mitochondrial enzyme carbamyl-phosphate synthetase I predisposes children to increased pulmonary artery pressure following surgical repair of congenital heart defects: a validated genetic association study

Increased pulmonary artery pressure (PAP) can complicate the postoperative care of children undergoing surgical repair of congenital heart defects. Endogenous NO regulates PAP and is derived from arginine supplied by the urea cycle. The rate-limiting step in the urea cycle is catalyzed by a mitochondrial enzyme, carbamoyl-phosphate synthetase I (CPSI). A well-characterized polymorphism in the gene encoding CPSI (T1405N) has previously been implicated in neonatal pulmonary hypertension. A consecutive modeling cohort of children (N=131) with congenital heart defects requiring surgery was prospectively evaluated to determine key factors associated with increased postoperative PAP, defined as a mean PAP>20 mmHg for at least 1h during the 48h following surgery measured by an indwelling pulmonary artery catheter. Multiple dimensionality reduction (MDR) was used to both internally validate observations and develop optimal two-variable through five-variable models that were tested prospectively in a validation cohort (N=41). Unconditional logistic regression analysis of the modeling cohort revealed that age (OR=0.92, p=0.01), CPSI T1405N genotype (AC vs. AA: OR=4.08, p=0.04, CC vs. AA: OR=5.96, p=0.01), and Down syndrome (OR=5.25, p=0.04) were independent predictors of this complex phenotype. MDR predicted that the best two-variable model consisted of age and CPSI T1405N genotype (p<0.001). This two-variable model correctly predicted 73% of the outcomes from the validation cohort. A five-variable model that added race, gender and Down's syndrome was not significantly better than the two-variable model. In conclusion, the CPSI T1405N genotype appears to be an important new factor in predicting susceptibility to increased PAP following surgical repair of congenital cardiac defects in children.     

Bacteriophage K1-5 encodes two different tail fiber proteins, allowing it to infect and replicate on both K1 and K5 strains of Escherichia coli

A virulent double-stranded DNA bacteriophage, Phi K1-5, has been isolated and found to be capable of infecting Escherichia coli strains that possess either the K1 or the K5 polysaccharide capsule. Electron micrographs show that the virion consists of a small icosohedral head with short tail spikes, similar to members of the Podoviridae family. DNA sequence analysis of the region encoding the tail fiber protein showed two open reading frames encoding previously characterized hydrolytic phage tail fiber proteins. The first is the K5 lyase protein gene of Phi K5, which allows this phage to specifically infect K5 E. coli strains. A second open reading frame encodes a protein almost identical in amino acid sequence to the N-acetylneuraminidase (endosialidase) protein of Phi K1E, which allows this phage to specifically infect K1 strains of E. coli. We provide experimental evidence that mature phage particles contain both tail fiber proteins, and mutational analysis indicates that each protein can be independently inactivated. A comparison of the tail gene regions of Phi K5, Phi K1E, and Phi K1-5 shows that the genes are arranged in a modular or cassette configuration and suggests that this family of phages can broaden host range by horizontal gene transfer.