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Two new cassane-type diterpenes, phangininoxys D and E (1 and 2), together with five known compounds were isolated from the seeds of Caesalpinia crista Linn. The structures of these compounds were elucidated by means of various spectroscopic analyses. All the isolated compounds were evaluated for cytotoxicity activities against HeLa, HT-29 and KB cell lines, and compound 7 showed moderate selective activities against KB cell line with an IC50 value of 17.1 μM.

Right atrial (RA) tachyarrhythmias are not rare in patients with congenital heart disease and a history of cardiac surgery. This study investigated the usefulness of a crista catheter for 3-dimensional electroanatomical mapping of RA tachyarrhythmias. We consecutively included 35 patients (age, 43.2 ± 15.6 years; 15 men) who underwent an electrophysiological study with 3-dimensional electroanatomical mapping for RA tachycardia or flutter. In 13 patients with atrial flutter, we recorded and compared the electrical sequence in the anterior and posterior portions of the RA lateral wall. We used a crista catheter as a mapping catheter for 3-dimensional mapping in 12 patients (crista group), a lasso catheter in 12 patients (lasso group), and an ablation catheter in 11 patients (ablation group). We compared the 3-dimensional mapping points, time, and speed (mapping points per minute) among the groups. Atrial flutter was confirmed as cavotricuspid isthmus-dependent in all patients whose two atrial electrical sequences were the same direction and as atypical (including scar-related and dual-loop) in all patients whose sequences were in the opposite direction. Mapping speed in the crista group was significantly faster than in the lasso and ablation groups: median (interquartile range) 44.0 (35.5–69.4) points/min, 23.7 (17.8–29.8) points/min, and 8.2 (4.8–11.0) points/min, respectively (p = 0.001). A crista catheter is useful for high-density 3-dimensional electroanatomical mapping of complex RA tachyarrhythmias. Comparison of the electrical sequences in the anterior and posterior portions of the RA lateral wall is helpful for differentiating between cavotricuspid isthmus-dependent and atypical atrial flutter.

SummaryThe mitochondrial contact site and cristae organizing system (MICOS) is a conserved multi-subunit complex crucial for maintaining the characteristic architecture of mitochondria. Studies with deletion mutants identified Mic10 and Mic60 as core subunits of MICOS. Mic60 has been studied in detail; however, topogenesis and function of Mic10 are unknown. We report that targeting of Mic10 to the mitochondrial inner membrane requires a positively charged internal loop, but no cleavable presequence. Both transmembrane segments of Mic10 carry a characteristic four-glycine motif, which has been found in the ring-forming rotor subunit of F1Fo-ATP synthases. Overexpression of Mic10 profoundly alters the architecture of the inner membrane independently of other MICOS components. The four-glycine motifs are dispensable for interaction of Mic10 with other MICOS subunits but are crucial for the formation of large Mic10 oligomers. Our studies identify a unique role of Mic10 oligomers in promoting the formation of inner membrane crista junctions.

Spontaneous discharge patterns of first-order canal afferents were analyzed in cats anethetized with pentobarbital sodium with particular emphasis on the relationship of regularity of resting discharge, sensitivity to angular acceleration and adaptation to the time delay between electrical labyrinthine stimulation and recording from afferents near Scarpa's ganglion. Regular units were found to have a high resting rate, low sensitivity to angular acceleration, were mostly nonadapting during prolonged acceleration and showed relatively long latency to electrical stimulation. Irregular units tended to have a low resting rate, high sensitivity, frequently showed adaptation and had short latencies. Intermediate neurons had mixed characteristics of regular and irregular units. In medulated nerve fibers, a direct relation exists between conduction velocity and fiber diameter. As latency is due primarily to conduction in the first-order axon, we may speculate that regular neurons have thin fibers which innervate the slope of the crista, irregular neurons have thick fibers which innervate the summit, and intermediate units have medium caliber fibers which innervate both the slope and summit of the crista ampullaris.

In the below paragraph we refer to Lysakowski et al. (2011) for the use of calbindin, while the right reference has to be Leonard and Kevetter (2002). In some experiments, an antibody for calbindin was also used to show the calyx nerve terminals (Leonard and Kevetter, 2002 instead of Lysakowski et al., 2011). Figure 6 shows such an example in a horizontal crista; note that calbindin antibody (red) clearly stained several afferent calyces, mostly located in the central zone, where GAD67 was conversely absent.

Previous in-vivo studies have shown that the tonotopically-organized hearing organ, the crista acustica, of the bushcricket Mecopoda elongata is a promising model for investigations of basic hearing mechanics. This linearly arranged organ with a length of

Glycosaminoglycans (GAGs) are ubiquitous in the inner ear, and disorders altering their structure or production often result in debilitating hearing and balance deficits. The specific mechanisms responsible for loss of hair-cell function are not well understood. We recently reported that introduction of a novel BODIPY conjugated xyloside (BX) into the endolymph primes fluorescent GAGs in vivo [6, 15]. Confocal and two-photon fluorescence imaging revealed rapid turnover and assembly of a glycocalyx enveloping the kinocilia and extending into the cupula, a structure that presumably serves as a mechanical link between the hair bundle and the cupula. Extracellular fluorescence was also observed around the basolateral surface of hair cells and surrounding afferent nerve projections into the crista. Single unit afferent recordings during mechanical hair bundle stimulation revealed temporary interruption of synaptic transmission following BX administration followed by recovery, demonstrating an essential role for GAGs in function of the hair cell synapse. In the present work we present a pharmacokinetic model to quantify the time course of BX primed GAG production and turnover in the ear.

The differential diagnosis of intracardiac masses that involves the right atrial chamber include vegetation, thrombus or tumours, and anatomic normal variants such as crista terminalis. Size, shape,location, mobility and attachment of the mass combined with the clinical findings help differentiate etiology. Two Dimensional Trans Thoracic Echocardiography (TTE) became the gold standard test for the diagnosis of intracardiac masses and further examination such as transesophagealechocardiography (TEE) and Cardiac Magnetic Resonance (CMR) improved the accuracy. The cardiovascular imaging is crucial to establish a correct diagnosis for proper management and therapy. Here the author describes the case report of a healthy middle-aged athlete who underwent echocardiography for sports preparticipation screening which detected the presence of a peduncolated right atrial mass of uncertain nature. Because of the strong suspicion for small right atrial mixoma, he underwent CMR that showed a small enhancing mass adherent to the right atrial posterior wall typical of prominent crista terminalis. This report of rare Crista Terminalis muscular bridge that mimicks right atrial mixoma gives new information for the understanding of the aspect of complex and strange right atrial anatomy

The high-frequency hearing organ of bushcrickets – the crista acustica (CA) – is tonotopically organized. Details about the mechano-electrical transduction mechanisms within the sensory-cell complex, however, remain unknown. In the recent study, we investigated and compared the anatomical, mechanical and electrophysiological properties of the CA and reveal a strong correlation of the mechanical and neuronal frequency tuning, which is supported by an anatomical gradient along the CA. Only in the distal high-frequency region of the CA a discrepancy between a strong mechanical response to low frequencies 30 kHz was found. Therefore, we suggest that there might be additional intrinsic tuning mechanisms in the sensory cells of the distal region to distinguish the frequency content of sound.

Energy requirements of aerobic cells are met by oxidation of carbohydrates, lipids and proteins which yields reduced coenzymes NADH and FADH 2 . The oxidation of NADH and FADH 2 proceeds in a highly organized system known as electron transport chain (ETC) which consists of complexes of proteins and coenzymes. The ultimate electron acceptor is oxygen, which is converted to water. The energy is conserved with the transport of electrons in ETC as electrochemical proton gradient, because protons are pumped out of the matrix to the intermembranal space. The electrochemical proton gradient drives the conversion of ADP to ATP, when protons are channeled back into the matrix through the ATP synthase. ETC is organized into four complexes I, II, III and IV. Each complex consists of unique species of molecules that include flavoproteins with tightly bound FAD or FMN, Coenzyme Q, cytochromes, iron–sulfur proteins, and copper bound proteins. The four complexes function as independent entities in a dynamic manner. The direction of electron flow in the ETC is from a higher energy to lower energy level.

The sensory patches of the vertebrate inner ear, which contain hair cells and supporting cells, are essential for hearing and balance functions. How the stereotypically organized sensory patches are formed remains to be determined. In this study, we isolated a zebrafish mutant in which the jag1b gene is disrupted by an EGFP insertion. Loss of Jag1b causes cell death in the developing posterior crista and results in downregulation of fgf10a in the posterior prosensory cells. Inhibition of FGFR activity in wild-type embryos also causes loss of the posterior crista, suggesting that Fgf10a mediates Jag1b activity. By contrast, in the anterior prosensory domain, Jag1b regulates separation of a single morphogenetic field into anterior and lateral cristae by flattening cells destined to form a nonsensory epithelium between the two cristae. MAPK activation in the nonsensory epithelium precursors is required for the separation. In the jag1b mutant, MAPK activation and cell flattening are extended to anterior crista primordia, causing loss of anterior crista. More importantly, inhibition of MAPK activity, which blocks the differentiation of nonsensory epithelial cells, generated a fused large crista and extra hair cells. Thus, Jag1b uses two distinct mechanisms to form three sensory cristae in zebrafish.

Interfibrillar mitochondria (IFM) of the heart in aged Fischer 344 rats show a biochemical defect which might be reflected in their morphology. We examined by high resolution scanning electron microscopy over 5500 mitochondria to determine if a concomitant structural alteration existed. This methodology provides a means of examining mitochondrial cristae in three dimensions. Cristae of in situ subsarcolemmal mitochondria (SSM) and of IFM in both 6- and 24-month-old Fischer rats are predominantly lamelliform. When isolated, these organelles, whether of SSM or IFM origin, display enhanced heterogeneity, but they have similar crista morphology irrespective of the age of the rat. Crista configuration does not play a major role in age-related cardiac mitochondrial defects.

SummaryMitochondrial outer- and inner-membrane fusion events are coupled in vivo but separable and mechanistically distinct in vitro, indicating that separate fusion machines exist in each membrane. Outer-membrane fusion requires trans interactions of the dynamin-related GTPase Fzo1, GTP hydrolysis, and an intact inner-membrane proton gradient. Inner-membrane fusion also requires GTP hydrolysis but distinctly requires an inner-membrane electrical potential. The protein machinery responsible for inner-membrane fusion is unknown. Here, we show that the conserved intermembrane-space dynamin-related GTPase Mgm1 is required to tether and fuse mitochondrial inner membranes. We observe an additional role of Mgm1 in inner-membrane dynamics, specifically in the maintenance of crista structures. We present evidence that trans Mgm1 interactions on opposing inner membranes function similarly to tether and fuse inner membranes as well as maintain crista structures and propose a model for how the mitochondrial dynamins function to facilitate fusion.

Carbon monoxide concentrations are derived from the CRyogenic Infrared Spectrometers and Telescopes for the Atmosphere (CRISTA) measurements in November 1994 and in August 1997. CRISTA is a helium-cooled infrared limb sounder. Its measurements include daytime spectra of the CO (Δ v =1) band at 4.7 μm. From the spectra CO densities are derived using a state of the art non-LTE model. During the first mission data were recorded between 58°S and about 68°N. In the second mission the latitude coverage was extended to ±71° in the mesosphere. The orbital characteristics, however, were such that daytime conditions prevailed in the northern (summer) hemisphere only. The derived CO concentrations agree well with other experimental data. As the CO mixing ratio increases steeply with altitude in the mesosphere and as the chemical lifetime is much longer than the time constants of dynamical motions this gas is an excellent tracer for dynamics. During the two CRISTA missions the CO distribution was found extremely variable both in time and in space, which is a result of the complex dynamics at these altitudes. The variations are compatible with the large-scale transport from the summer to the winter hemisphere. The influence of planetary waves is observed under winter as well as under summer conditions. Large tidal amplitudes were found at mid-latitudes during the second CRISTA mission. The overall wave activity observed in August by CRISTA suggests an abrupt change of wave damping in the vicinity of 97 km.

There is increasing evidence now that F(1)F(0) ATP synthase is arranged in dimers in the inner mitochondrial membrane of several organisms. The dimers are also considered to be the building blocks of oligomers. It was recently found that the monomers in beef and the alga Polytomella ATP synthase dimer make an angle of approximately 40 degrees and approximately 70 degrees, respectively. This arrangement is considered to induce a strong local bending of the membrane. To further understand the packing of dimers into oligomers we performed an electron microscopy analysis of ATP synthase dimers purified from Saccharomyces cerevisiae. Two types of dimers were found in which the angle between the monomers is either approximately 90 degrees or approximately 35 degrees. According to our interpretation, the wide-angle dimers (70-90 degrees) are "true-dimers" whereas the small-angle dimers (35-40 degrees) rather are "pseudo-dimers", which represent breakdown products of two adjacent true dimers in the oligomer. Ultrathin sectioning of intact Polytomella mitochondria indicates that the inner mitochondrial or cristae membrane is folded into lamellae and tubuli. Oligomers of ATP synthase can arrange in a helical fashion in tubular-shaped cristae membranes. These results strongly support the hypothesized role of ATP synthase oligomers in structural determination of the mitochondrial inner membrane.

A new dimer 1 and two new cassane-type diterpenes 2 and 3, designated taepeenin J–L, were isolated from the seeds of Caesalpinia crista L. Compound 1 possesses a dimeric vouacapane skeleton. Their structures were elucidated on the basis of spectroscopic analysis.

Synaptic activity was recorded with sharp microelectrodes during rest and during 0.3-Hz sinusoidal stimulation from bouton afferents identified by their efferent-mediated inhibitory responses. A glutamate antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) decreased quantal size (qsize) while lowering external Ca(2+) decreased quantal rate (qrate). Miniature excitatory postsynaptic potentials (mEPSPs) had effective durations (qdur) of 3.5-5 ms. Their timing was consistent with Poisson statistics. Mean qsizes ranged in different units from 0.25 to 0.73 mV and mean qrates from 200 to 1,500/s; there was an inverse relation across the afferent population between qrate and qsize. qsize distributions were consistent with the independent release of variable-sized quanta. Channel noise, measured during AMPA-induced depolarizations, was small compared with quantal noise. Excitatory responses were larger than inhibitory responses. Peak qrates, which could approach 3,000/s, led peak excitatory mechanical stimulation by 40 degrees . Quantal parameters varied with stimulation phase with qdur and qsize being maximal during inhibitory stimulation. Voltage modulation (vmod) was in phase with qrate and had a peak depolarization of 1.5-3 mV. On average, 80% of vmod was accounted for by quantal activity; the remaining 20% was a nonquantal component that persisted in the absence of quantal activity. The extracellular accumulation of glutamate and K(+) are potential sources of nonquantal transmission and may provide a basis for the inverse relation between qrate and qsize. Comparison of the phases of synaptic and spike activity suggests that both presynaptic and postsynaptic mechanisms contribute to variations across afferents in the timing of spikes during sinusoidal stimulation.

—Nine new cassane-type diterpenes, named taepeenin A–I, and two new norcassane-type diterpenes, named nortaepeenin A–B,were isolated from the stems and roots of Caesalpinia crista along with three known diterpenes: vinhaticoic acid, methyl vinhaticoate andent-11b-hydroxy-rosa-5,15-diene. Their structures were elucidated on the basis of spectroscopic analysis. In addition, the structure oftaepeenin A was confirmed by X-ray diffraction analysis.q 2005 Elsevier Ltd. All rights reserved. 1. IntroductionCassane-type diterpenoids are characteristic components ofthe genus Caesalpinia of the Leguminosae family. One ofthem, the plant Caesalpinia crista L., known locally as‘Taepee’ in Thai, is a climber distributed from India andCeylon through most of Southeast Asia to the Ryu-KyuIslands, Queensland, and Caledonia. 1 The leaves, roots, andfruits of this plant are used as a tonic and an antiperiodic. 2 The seed kernels of C. crista displayed interestingantimalarial activity. 3 Previous chemical investigations ofthe seed kernels have revealed the presence of severalcassane-type diterpenoids.

Most information on the properties of mammalian vestibular primary afferents has been obtained in deeply anesthetized animals, in vivo. Generally, non-human primates and larger rodents have been the species of choice. Investigations using smaller rodents, such as the laboratory mouse, have been limited despite the increasing availability of naturally occurring or engineered mutants that result in balance disorders. Furthermore, in vitro preparations of the intact peripheral vestibular apparatus are only available for non-mammalian vertebrates. To take advantage of the genetic/molecular advances available in mice and the utility of in vitro preparations that permit manipulations of the extracellular milieu, we developed an isolated mouse inner ear preparation with the attached eighth cranial nerve for electrophysiological recording. Intra-axonal recordings of background activity in vestibular primary afferents were obtained in a modified Ringer's solution (0.25 mM Ca2+; 3.25 mM Mg2+) at 22 °C. We also recorded afferent activity in the presence of neuroactive drugs known to affect various stages of the transduction cascade. These results, together with responses to sinusoidal mechanical deformation of the membranous ducts, showed that transduction mechanisms remain viable. Where possible, we also obtained results in vivo for comparison. In future, the in vitro mouse preparation will allow investigation of the effects of genetic manipulations and pharmacological agents on the intact peripheral vestibular apparatus.

Introduction: Although the role of action potential duration restitution (APD-R) in the initiation and maintenance of ventricular fibrillation (VF) has been the subject of numerous investigations, its role in the generation of atrial fibrillation (AF) is less well studied. The cellular and ionic basis for coarse versus fine AF is not well delineated. Methods and Results: We measured APD-R during acetylcholine-mediated AF as well as during pacing (standard and dynamic protocols) in crista teriminalis, pectinate muscle, superior vena cava, and appendage of isolated canine arterially perfused right atria (n = 15). Transmembrane action potential (TAP), pseudo-ECG, and isometric tension development were simultaneously recorded. Acetylcholine flattened APD-R measured by both standard and dynamic protocols, but promoted induction of AF. AF was initially coarse, converting to fine within 3–15 minutes of AF. Coarse, but not fine AF was associated with dramatic fluctuations in tension development, reflecting wide variations in intracellular calcium activity ([Ca2+]i). During coarse AF, APD-R data displayed a cloud-like distribution pattern, with a wide range of maximum APD-R slope (from 1.21 to 0.35). A maximum APD-R slope >1 was observed only in crista terminalis (3/10). The APD-R relationship was relatively linear and flat during fine AF. Reduction of [Ca2+]i was associated with fine AF whereas augmentation of [Ca2+]i with coarse AF. Conclusions: Our data indicate that while APD-R may have a limited role in the maintenance of coarse AF, it is unlikely to contribute to the maintenance of fine AF and that [Ca2+]i dynamics determine the degree to which AF is coarse or fine.

Objective The aim of this study is to determine the immediate effects of intraperitoneal doses of gentamicin (GM) which would result in variable degrees of destruction of crista ampullary hair cells of frogs. This information will serve as a baseline guide to cell regeneration experiments on the damaged vestibular sense organ. Study design The American bullfrog was administered daily intraperitoneal doses of 50, 100, 150, and 200 mg/kg of GM for 7 days. Animals were sacrificed 1 day after the final injection for cytomorphic evaluation. Histologically processed posterior semicircular duct cristae were resin-embedded, and tissue samples were subjected to serial cross sectioning of the crista from the periphery to the central zone using glass knives in an ultramicrotome. Stained sections were analyzed in light microscope using ocular grid micrometry. The areal density of nuclear profiles of the vestibular sensory and supporting cells (sensory cells [SNCs] and supporting cells [SPCs], respectively; number per square millimeter) and the nuclear diameter of SNCs were manually determined. Results A 7-day administration of GM produced noticeable quantitative alteration of the posterior crista hair cells and SPCs. Histological analysis revealed a significant decrease in the density of SNCs and a concomitant increase in the density of SPCs (1-way analysis of variance). Conclusion and significance The cytomorphic data derived from this study show that 4 doses of intraperitoneal gentamicin administered to the bullfrog caused a decline in the areal density of sensory hair cells of the posterior canal crista ampullaris. Also noted was an increase in the density of adjacent SPCs. Although speculative, the increase in SPC population could be a harbinger of regeneration of the vestibular hair cells as suggested by other investigators in different species. The significance of present observations will be helpful to initiate future studies related to recovery of SNCs in a similarly damaged frog ampullary organ. Through a standardized quantitative approach to the study of SNCs and SPCs of the crista organ, the vestibulo-toxicity of newly developed drugs can be assessed.

During the CRISTA-1 mission three pronounced fingerlike structures reaching from the lower latitudes to the mid-latitudes, so-called streamers, were observed in the measurements of several trace gases in early November 1994. A simulation of these streamers in previous studies employing the KASIMA (Karlsruhe Simulation Model of the Middle Atmosphere) and ROSE (Research on Ozone in the Stratosphere and its Evolution) model, both being Eulerian models, show that their formation is due to adiabatic transport processes. Here, the impact of mixing on the development of these streamers is investigated. These streamers were simulated with the CLaMS model (Chemical Lagrangian Model of the Stratosphere), a Lagrangian model, using N2O as long-lived tracer. Using several different initialisations the results were compared to the KASIMA simulations and CRISTA (Cryogenic Infrared Spectrometer and Telescope for the Atmosphere) observations. Further, since the KASIMA model was employed to derive a 9-year climatology, the quality of the reproduction of streamers from such a study was tested by the comparison of the KASIMA results with CLaMS and CRISTA. The streamers are reproduced well for the Northern Hemisphere in the simulations of CLaMS and KASIMA for the 6 November 1994. However, in the CLaMS simulation a stronger filamentation is found while larger discrepancies between KASIMA and CRISTA were found especially for the Southern Hemisphere. Further, compared to the CRISTA observations the mixing ratios of N2O are in general underestimated in the KASIMA simulations. An improvement of the simulations with KASIMA was obtained for a simulation time according to the length of the CLaMS simulation. To quantify the differences between the simulations with CLaMS and KASIMA, and the CRISTA observations, the probability density function technique (PDF) is used to interpret the tracer distributions. While in the PDF of the KASIMA simulation the small scale structures observed by CRISTA are smoothed out due to the numerical diffusion in the model, the PDFs derived from CRISTA observations can be reproduced by CLaMS by optimising the mixing parameterisation. Further, this procedure gives information on small-scale variabilities not resolved by the CRISTA observations.

An along-track analysis of temperature height profiles measured by the CRISTA satellite instrument was performed to obtain height profiles of absolute values of gravity wave momentum flux. Global distributions of momentum flux from CRISTA are compared to results of the Warner and McIntyre gravity wave parameterization scheme. Only a limited range of horizontal and vertical wavelengths is visible to the CRISTA instrument. Therefore the model results have to be filtered accordingly to make comparisons meaningful. We demonstrate the importance of such a wavelength filtering which significantly increases the correlation of observed and modeled momentum fluxes.

Seven new cassane-type diterpenes, caesalpinin MF-ML (1-7), and a new norcassane-type diterpene, norcaesalpinin MD (8), have been isolated from the CH2Cl2 extract of seed kernels of Caesalpinia crista from Myanmar, together with sixteen known cassane-type diterpenes, 7-acetoxybonducellpin C, caesaldekarin e, caesalmin C, caesalmin G, 2-acetoxycaesaldekarin e, zeta-caesalpin, caesalpinin D, caesalpinin E, caesalpinin F, caesalpinin H, caesalpinin I, caesalpinin J, caesalpinin K, caesalpinin M, caesalpinin N, and caesalpinin O. The structures of the isolated compounds were elucidated by the use of spectroscopic techniques.

Respiration in all cells depends upon synthesis of ATP by the ATP synthase complex, a rotary motor enzyme. The structure of the catalytic moiety of ATP synthase, the so-called F(1) headpiece, is well established. F(1) is connected to the membrane-bound and ion translocating F(0) subcomplex by a central stalk. A peripheral stalk, or stator, prevents futile rotation of the headpiece during catalysis. Although the enzyme functions as a monomer, several lines of evidence have recently suggested that monomeric ATP synthase complexes might interact to form a dimeric supercomplex in mitochondria. However, due to its fragility, the structure of ATP synthase dimers has so far not been precisely defined for any organism. Here we report the purification of a stable dimeric ATP synthase supercomplex, using mitochondria of the alga Polytomella. Structural analysis by electron microscopy and single particle analysis revealed that dimer formation is based on specific interaction of the F(0) parts, not the F(1) headpieces which are not at all in close proximity. Remarkably, the angle between the two F(0) part is about 70 degrees, which induces a strong local bending of the membrane. Hence, the function of ATP synthase dimerisation is to control the unique architecture of the mitochondrial inner membrane.

Whole membranous labyrinths of the bull frogs were used in order to replicate the human vestibule.The posterior semicircular canals (PSC) were exposed leaving the remaining membranous labyrinth encapsulated in the otic capsule. The membranous labyrinth was cut by 0.5 mm at the crus commune to create a tiny opening. The cupula was visualized by gently injecting India ink of the opening. The motion of the cupula was analyzed during compression of the canal wall by a needle. The motion appeared to be that of a "diaphragma" when the compression was weak. The motion appeared to be that of a "swing door" with the base of the cupula hinged on the crista when the compression was strong.A small piece of otoconia removed from the sacculus of the other ear was introduced through this opening into the canal lumen. The position of the preparation was controlled so that the otoconia were attached onto the cupular surface. This was regarded as a cupulolithiasis model. We performed the same experiments on the cupulolithiasis models and results were almost same, except that the motion of the cupula of cupulolithiasis models was shorter than those of the normal models. It was inferred that the motion of the cupula of cupulolithiasis models was suppressed by the weight of the otoconia.It was observed that the india ink traversed the cupula during the compression and even the otoconia traversed the cupula. Therefore, it was inferred that the adhesion of the cupula and the ampula wall is not strong, thus protecting the cupula from detaching from the crista.

Noise over-stimulation will induce or influence molecular pathways in the cochlea; one approach to the identification of the components of these pathways in the cochlea is to examine genes and proteins that change following different types and levels of stress. Quantitative reverse transcription polymerase chain reaction provides a method to look at differential expression of genes in the acoustic stress response. By using this technique we have revealed a down-regulation of the level of otospiralin mRNA in the cochlea of guinea pigs after white noise over-stimulation for 2 h at 108 dB SPL. Otospiralin represents an inner ear specific protein found in fibrocytes of spiral limbus and spiral ligament in the cochlea, and some regions of the vestibule as the stroma underlying the utricle and crista sensory epithelia and the subepithelial layer of the walls of semicircular canals and maculae. It has been recently reported that transient down-regulation of otospiralin in guinea pigs causes vestibular syndrome and deafness. Our results suggest a possible role of this gene in response to acoustical stress, although the exact mechanism remains to be resolved.

The aim of this study is to determine the type of skull as well as to examine its internal appearance and configuration of skull base. Special attention is given to the direction and position of the pyramid of the temporal bone, the volume and appearance of all the three cranial fossas, and the direction and appearance of crista alaris - all of those compared to the skull type.Considering the obtained results (specially for crista alaris and middle fossa) we can ascertain outstanding independence of sphenoid bone and its parts in the formation of cranial base. Located in the middle of the skull, in front of strong pyramids transversal axis and two vertical axis, frontal crest and internal occipital crest, it by itself with her lesser wing presents an important transversal axis of cranial base. Cases in which crista alaris with its position does not follow the type of skull (in 20 % cases crista alaris does not follow skull protuberances), may probably be explained by strong and independent development of this bone, which is placed among other bones of cranial base like a peg. It also, by itself dictates form and configuration of the middle part of skull. That is also confirmed by middle fossa which, according to its position in the middle of cranial base and relation with sphenoid bone, shows significant deviation with respect to posterior fossa (follows the type of skull in 47 % cases).

The present study was conducted to visualize the ultrastructural features of vestibular efferent boutons in the oyster toadfish, Opsanus tau. The crista ampullaris of the horizontal semicircular canal was processed for and examined by routine transmission electron microscopy. The results demonstrate that such boutons vary in size and shape, and contain a heterogeneous population of lucent vesicles with scattered dense core vesicles. Efferent contacts with hair cells are characterized by local vesicle accumulations in the presynaptic terminal and a subsynaptic cistern in the postsynaptic region of the hair cell. Serial efferent to hair cell to afferent synaptic arrangements are common, particularly in the central portion of the crista. However, direct contacts between efferent terminals and afferent neurites were not observed in our specimens. The existence of serial synaptic contacts, often with a row of vesicles in the efferent boutons lining the efferent-afferent membrane apposition, suggests that the efferent influence on the crista may involve both synaptic and nonsynaptic, secretory mechanisms. Further, it is suggested that differences in more subtle aspects of synaptic architecture and/or transmitter and receptor localization and interaction may render the efferent innervation of the peripheral crista less effective in influencing sensory processing.

Although γ-aminobutyric acid (GABA) and glutamate are known to be present in the vestibular sensory epithelia of a variety of species, the functional relationship between these two transmitters is not clear. The present study addresses the three-dimensional spatial distribution of GABA and glutamate immunoreactivity in the vestibular labyrinth of the oyster toadfish by using whole end organs labeled by immunofluorescence with monoclonal anti-GABA and/or antiglutamate antibodies and visualized as whole mounts by multiphoton confocal microscopy. We find glutamate-immunoreactive hair cells present throughout the sensory epithelium. In contrast, prominent GABA immunoreactivity is restricted to a small population of hair cells located in the central region of the crista. Double immunofluorescence reveals two distinct staining patterns in GABA-labeled hair cells. Most (∼80%) GABA-labeled cells show trace levels of glutamate, appropriate for the metabolic/synthetic role of cytoplasmic glutamate. The remainder of the GABA-stained cells contain substantial levels of both GABA and glutamate, suggesting transmitter colocalization. In the toadfish utricle, glutamatergic hair cells are present throughout the macula. GABA-immunoreactive hair cells follow the arc of the striola, and most GABA-labeled receptor cells coexpress glutamate. The localization of GABA was explored in other species as well. In the pigeon, GABAergic hair cells are present throughout the crista ampullaris. Our findings demonstrate that multiple, neurochemically distinct types of hair cells are present in vestibular sensory epithelia. These observations, together with the excitatory activity generally associated with 8th nerve afferent fibers, strongly suggest that GABA serves an important, specific, and complex role in determining primary afferent response dynamics. J. Comp. Neurol. 471:1–10, 2004. © 2004 Wiley-Liss, Inc.

Semiquantitative histological evaluation was performed on inner ears from Mongolian gerbils to study vestibular damages using ototoxic drugs. Comparisons were made between animals receiving five daily injections of gentamicin/gelfoam slurry, streptomycin/gelfoam slurry, saline/gelfoam slurry and noninjected controls. For gentamicin treated ears, when percent decrease of nuclei in sensory cell layer was compared, the damage to the sensory cells was significantly more severe in the posterior crista than in the superior and lateral crista from two of three gerbils. For streptomycin treated ears, when percent decrease of nuclei in sensory cell layer was compared, the damage to the posterior crista was similar to superior and lateral crista. These results indicate that our models are useful for detecting different vestibulotoxicity among three cristae.

The vertebrate inner ear is a marvel of structural and functional complexity, which is all the more remarkable because it develops from such a simple structure, the otic placode. Analysis of inner ear development has long been a fascination of experimental embryologists, who sought to understand cellular mechanisms of otic placode induction. More recently, however, molecular and genetic approaches have made the inner ear a useful model system for studying a much broader range of basic developmental mechanisms, including cell fate specification and differentiation, axial patterning, epithelial morphogenesis, cytoskeletal dynamics, stem cell biology, neurobiology, physiology, etc. Of course, there has also been tremendous progress in understanding the functions and processes peculiar to the inner ear. The goal of this review is to recount how historical approaches have shaped our understanding of the signaling interactions controlling early otic development; to discuss how new findings have led to fundamental new insights; and to point out new problems that need to be resolved in future research.

When studied in vitro, type I hair cells in amniote vestibular organs have a large, negatively activating K+ conductance. In type II hair cells, as in nonvestibular hair cells, outwardly rectifying K+ conductances are smaller and more positively activating. As a result, type I cells have more negative resting potentials and smaller input resistances than do type II cells; large inward currents fail to depolarize type I cells above –60 mV. In nonvestibular hair cells, afferent transmission is mediated by voltage-gated Ca2+ channels that activate positive to –60 mV. We investigated whether Ca2+ channels in type I cells activate more negatively so that quantal transmission can occur near the reported resting potentials. We used the perforated patch method to record Ca2+ channel currents from type I and type II hair cells isolated from the rat anterior crista (postnatal days 4–20). The activation range of the Ca2+ currents of type I hair cells differed only slightly from that of type II cells or nonvestibular hair cells. In 5 mM external Ca2+, currents in type I and type II cells were half-maximal at –41.1 ± 0.5 (SE) mV ( n = 10) and –37.2 ± 0.2 mV ( n = 10), respectively. In physiological external Ca2+ (1.3 mM), currents in type I cells were half-maximal at –46 ± 1 mV ( n = 8) and just 1% of maximal at –72 mV. These results lend credence to suggestions that type I cells have more positive resting potentials in vivo, possibly through K+ accumulation in the synaptic cleft or inhibition of the large K+ conductance. Ca2+ channel kinetics were also unremarkable; in both type I and type II cells, the currents activated and deactivated rapidly and inactivated only slowly and modestly even at large depolarizations. The Ca2+ current included an L-type component with relatively low sensitivity to dihydropyridine antagonists, consistent with the α subunit being CaV1.3 (α1D). Rat vestibular epithelia and ganglia were probed for L-type α-subunit expression with the reverse transcription-polymerase chain reaction. The epithelia expressed CaV1.3 and the ganglia expressed CaV1.2 (α1C).

The pathophysiology of benign paroxysmal positional vertigo (BPPV) is not completely understood. Although the concept of degenerated otoconia transforming the posterior canal (PC) crista into a gravity-sensitive sense organ has gained popular support, several temporal bone (TB) series have revealed similar deposits in normal TBs, suggesting they are a normal change in the aging labyrinth. Furthermore, some TBs from patients with BPPV do not contain particles in the posterior canal. Five TBs from patients with BPPV were studied quantitatively and qualitatively. A small PC cupular deposit was found in 1 TB, while none was seen in the other 4 TBs. The major pathological changes were 1) a 50% loss of ganglion cells in the superior vestibular division of all 5 TBs and 2) a 50% loss of neurons in the inferior division of 3 TBs, and a 30% loss in 2 TBs that contained abnormal saccular ganglion cells. These observations support a concept in the pathophysiology of BPPV that includes loss of the inhibitory effect of otolith organs on canal sense organs.

The inner membrane system of mitochondria is known to consist of two contiguous but distinct membranes: the inner boundary membrane, which apposes the outer membrane, and the cristal membrane, which forms tubules or lamellae in the interior. Using immunolabeling and transmission electron microscopy of bovine heart tissue, we have calculated that around 94% of both Complex III of the respiratory chain and the ATP synthase are located in the cristal membrane, and only around 6% of either is in the inner boundary membrane. When accounting for the topographical ratio of cristal membrane versus inner boundary membrane, we find that both complexes exist at a 2.2–2.6-fold higher concentration in the cristal membrane. The residual protein in the inner boundary membrane may be newly assembled complexes destined for cristal membranes. Our results argue for restricted diffusion of complexes through the cristal junctions and indicate that the mitochondrial cristae comprise a regulated submitochondrial compartment specialized for ATP production.

The simple primordium of the inner ear (otocyst) differentiates into many cell types, including sensory neurons and hair cells. We examined expression of the divergent homeobox transcription factor, cProx1, during otocyst development in chickens. Nuclear cProx1 protein is not evident in the otic placode but emerges in the otic cup by stage 12. At stage 16, cProx1-positive nuclei are scattered continuously throughout the neuroepithelium, from anteroventral to posteromedial. These labeled cells are neural precursors; they express betaIII-tubulin and migrate to the cochleovestibular ganglion between stages 13 and 21. By stage 18, two areas develop a dense pattern of cProx1 expression in which every nucleus is labeled. These areas emerge at the anterior and posterior extremes of the band of scattered cProx1 expression and express the sensory markers cSerrate1 and Cath1 by stage 23. Four discrete patches of dense cProx1 expression appear by stage 23 that correspond to the future superior crista, lateral crista, saccular macula, and posterior crista, as confirmed by immunolabeling for hair cell antigen (HCA) by stage 29. The remaining sensory epithelia display a dense pattern of cProx1 expression and label for HCA by stage 29. In the basilar papilla, nuclear cProx1 expression is down-regulated in most hair cells by stage 37 and in many supporting cells by stage 40. Our findings show that regions of the otocyst that give rise to neurons or hair cells are distinguished by their relative density of cProx1-positive nuclei, and suggest a role for cProx1 in the genesis of these cell types.

The present study describes the localization and distribution of putative ecto-nucleoside-triphosphate-diphosphohydrolases in the frog semicircular canals. These enzymes provide the terminating mechanism of adenosine-5'-triphosphate (ATP) signalling. The localization of the ATP hydrolysis was mapped ultracytochemically using a one-step cerium citrate reaction. Electron-dense precipitates, indicating ecto-adenosine-triphosphatase (ecto-ATPase) activity, were found at the outer surface of plasma membranes of crista hair cells and supporting cells of the sensory epithelium, transitional cells and undifferentiated cells of the ampullar wall and dark cells constituting the secretory epithelium. Non-sensory cells of the ampulla usually exhibited reaction deposits at the level of both apical and basolateral membranes coming into contact with the endolymph and the perilymph respectively, while cells constituting the sensory epithelium showed evident differences in relation to their position. Hair cells and supporting cells of the peripheral regions exhibited clear reaction products both at the level of apical and basolateral membranes, while those of the isthmus region showed abundant reactivity only at the level of their apical membranes. Of particular interest was the observation that hair cell stereocilia exhibited an abundant ecto-ATPase activity, thus suggesting a possible colocalization of enzymatic sites with purinergic receptors and mechanotransduction channels. This strategic expression of ecto-ATPase sites could provide a rapid mechanism of ATP removal able to rapidly restore the sensitivity of transduction channels. In conclusion, the widespread distribution of ecto-ATPase sites at the level of sensory and non-sensory cells of the frog semicircular canals suggests that ATP may have a key role in controlling vestibular function.

The projection of the posterior canal crista and saccular afferents to the cerebellum of embryonic and neonatal mice was investigated using carbocyanine dyes. Anterograde tracing from these two endorgans reveals a partial segregation of these two sets of afferents. The saccule projects predominantly to the uvula, with very minor input to the nodulus. The posterior canal projects mainly to the nodulus and, to a lesser extent, to the uvula. Retrograde tracing from the uvula and nodulus confirms this partial segregation for these two endorgans and extends it to other vestibular endorgans. Uvular injections result in many more labeled fibers in the gravistatic maculae than in the canals' cristae. In contrast, nodular injection reveals many more labeled fibers in the canal cristae than in the gravistatic maculae. This partial segregation may play a role in the information processing in these folia. Our developmental data suggest that the initial segregation at E17 coincides with the formation of the postero-lateral fissure. This embryonic segregation of the primary vestibular mossy fibers to the uvula and nodulus commences long before the maturity of their targets, the granule cells and unipolar brush cells. Thus, the segregation of the primary vestibular projection to the uvula and nodulus does not depend on cues related to the target cells. Rather, the segregation may reflect more global cerebellar patterning mechanisms involving guidance for the vestibular afferent fibers independent of the future target cells.

The turtle posterior crista is made up of two hemicristae, each consisting of a central zone containing type I and type II hair cells and a surrounding peripheral zone containing only type II hair cells and extending from the planum semilunatum to the nonsensory torus. Afferents from various regions of a hemicrista differ in their discharge properties. To see if afferent diversity is related to the basolateral currents of the hair cells innervated, we selectively harvested type I and II hair cells from the central zone and type II hair cells from two parts of the peripheral zone, one near the planum and the other near the torus. Voltage-dependent currents were studied with the whole cell, ruptured-patch method and characterized in voltage-clamp mode. We found regional differences in both outwardly and inwardly rectifying voltage-sensitive currents. As in birds and mammals, type I hair cells have a distinctive outwardly rectifying current ( IK,L), which begins activating at more hyperpolarized voltages than do the outward currents of type II hair cells. Activation of IK,Lis slow and sigmoidal. Maximal outward conductances are large. Outward currents in type II cells vary in their activation kinetics. Cells with fast kinetics are associated with small conductances and with partial inactivation during 200-ms depolarizing voltage steps. Almost all type II cells in the peripheral zone and many in the central zone have fast kinetics. Some type II cells in the central zone have large outward currents with slow kinetics and little inactivation. Although these currents resemble IK,L, they can be distinguished from the latter both electrophysiologically and pharmacologically. There are two varieties of inwardly rectifying currents in type II hair cells: activation of IK1is rapid and monoexponential, whereas that of Ihis slow and sigmoidal. Many type II cells either have both inward currents or only have IK1; very few cells only have Ih. Inward currents are less conspicuous in type I cells. Type II cells near the torus have smaller outwardly rectifying currents and larger inwardly rectifying currents than those near the planum, but the differences are too small to account for variations in discharge properties of bouton afferents innervating the two regions of the peripheral zone. The large outward conductances seen in central cells, by lowering impedances, may contribute to the low rotational gains of some central-zone afferents.

Rats were exposed to a hypergravity (HG) level of 2.5 x g from conception until the age of 14 weeks. The vestibular epithelia of four of these animals and four control animals were immunohistochemically labeled for actin and tubulin. The apical cross-sectional area of epithelial cells of HG exposed rats appeared to be larger in all end organs. Area increase was 7.0% in the utricle (p

In recent years, electron tomography has provided detailed three-dimensional models of mitochondria that have redefined our concept of mitochondrial structure. The models reveal an inner membrane consisting of two components, the inner boundary membrane (IBM) closely apposed to the outer membrane and the cristae membrane that projects into the matrix compartment. These two components are connected by tubular structures of relatively uniform size called crista junctions. The distribution of crista junction sizes and shapes is predicted by a thermodynamic model based upon the energy of membrane bending, but proteins likely also play a role in determining the conformation of the inner membrane. Results of structural studies of mitochondria during apoptosis demonstrate that cytochrome c is released without detectable disruption of the outer membrane or extensive swelling of the mitochondrial matrix, suggesting the formation of an outer membrane pore large enough to allow passage of holo-cytochrome c. The possible compartmentation of inner membrane function between the IBM and the cristae membrane is also discussed.

Vascular change may contribute to age-related vestibular dysfunction. Previously, we reported a significant age-related decrease in blood flow (BF) and mean capillary diameter (D(cap)) in the rat posterior canal crista. The purpose of this study was to examine an otolith organ, the utricle, for similar changes. Old male Fischer 344 rats (O; 28-31 mos) were anesthetized, and the left cardiac ventricle was transcutaneously injected with radioactive microspheres to determine BF. The temporal bones were removed, fixed, and decalcified. The utricles were dissected free and placed into a gamma counter with the reference samples. The specimens were then plastic embedded and serially sectioned at 1 microm according to the vertical section technique. Microsphere surface counts were made and neuroepithelial BF calculated. A systematic random set of sections was sampled and analyzed using stereological techniques for estimates of D(cap), capillary surface area/unit volume (S(v,cap)), capillary length/ unit volume (L(v,cap)), and volume of utricular neuroepithelium (V(ut)). Using these data, total capillary surface (S(cap)) and total length (L(cap)) were calculated. Statistical comparisons were made with data from our previous study of young animals (Y; 3-6 mos). Results indicate a significant age-related decrease in BF (Y = 0.125 microL/min, O = 0.062 microL/min; P = 0.003), D(cap) (Y = 5.95 micro, O = 4.57 microm; P = 0.0002), S(vcap) (Y = 12.33 mm2/mm3, = 9.87 mm2/mm3, P = 0.016), S(cap) (Y = 0.178 mm2, O = 0.129 mm2; p = 0.01), and V(ut) (Y = 0.014 mm3, O = 0.013 mm3; P = 0.04) with no significant change in L(v,cap) (Y = 655 mm/mm3, O = 686 mm/mm3, P = 0.41) or L(cap) (Y = 9.47 mm, O = 8.96 mm; P = 0.49). These age-related vascular changes are likely to have a significant impact on utricular physiological and thus, dysequilibrium.

The developmental segregation of gravistatic input mediated by saccular fibers and of angular acceleration input mediated by posterior crista (PC) fibers was analyzed for the first time in a developing mammal using carbocyanine dye tracing in fixed tissue. The data reveal a more extensive projection of either endorgan in 7-day-old mice (P7) than has previously been reported in adult mammals. While we confirm and extend many previous findings, we also describe a novel segregation of saccular and posterior crista fibers in the anterior half of the medial vestibular nucleus (Mv) not reported before. Our developmental analysis shows a progressive segregation of posterior crista and saccular fibers to their respective discrete projection areas between embryonic day 15 (E15) and birth (P0). Retention of overlap in young adult animals appears to reflect the early embryonic overlap found in most areas. The vestibular projection does not show a topological projection as has been described in many other sensory systems. We propose that the unique projection features of the vestibular endorgans may relate to the transformation of vestibular signals into a motor output in the three neuron reflex arc of the VOR, of which the primary vestibular projection constitutes the first leg.

The use of electron tomography has allowed the three-dimensional membrane topography of the mitochondrion to be better understood. The most striking feature of this topology is the crista junction, a structure that may serve to divide functionally the inner membrane and intermembrane spaces. In situ these junctions seem to have a preferred size and shape independent of the source of the mitochondrion with few exceptions. When mitochondria are isolated and have a condensed matrix the crista junctions enlarge and become nondiscrete. Upon permeation of the inner membrane and subsequent swelling of the matrix space, the uniform circular nature of the crista junction reappears. We examine the distribution of shapes and sizes of crista junctions and suggest a thermodynamic model that explains the distribution based on current theories of bilayer membrane shapes. The theory of spontaneous curvature shows the circular junction to be a thermodynamically stable structure whose size and shape is influenced by the relative volume of the matrix. We conclude that the crista junction exists predominantly as a circular junction, with other shapes as exceptions made possible by specific characteristics of the lipid bilayer.

The effect of prolonged exposure to hypergravity on the morphology of vestibular epithelia of rats was investigated. At the age of 1 month, i.e., when vestibular end organs are fully maturated, three rats were transferred to a hypergravity environment of 2.5 g inside a large radius centrifuge. After 9 months, vestibular epithelia of these animals and of three control animals were immunohistochemically labeled for actin and tubulin. The apical cross-sectional area of epithelial cells of hypergravity exposed rats appeared to be smaller in all end organs. Area reduction was 1.9% in the saccule (not significant), 5.0% in the utricle (p < 0.005), and 11.6% in the crista (p<

The inner membrane of the mitochondrion folds inwards, forming the cristae. This folding allows a greater amount of membrane to be packed into the mitochondrion. The data in this study demonstrate that subunits e and g of the mitochondrial ATP synthase are involved in generating mitochondrial cristae morphology. These two subunits are non‐essential components of ATP synthase and are required for the dimerization and oligomerization of ATP synthase. Mitochondria of yeast cells deficient in either subunits e or g were found to have numerous digitations and onion‐like structures that correspond to an uncontrolled biogenesis and/or folding of the inner mitochondrial membrane. The present data show that there is a link between dimerization of the mitochondrial ATP synthase and cristae morphology. A model is proposed of the assembly of ATP synthase dimers, taking into account the oligomerization of the yeast enzyme and earlier data on the ultrastructure of mitochondrial cristae, which suggests that the association of ATP synthase dimers is involved in the control of the biogenesis of the inner mitochondrial membrane.

Mitochondrial membrane architecture is important for organelle function. Alterations thereof are linked to a number of human disorders including diabetes and cardiomyopathy. The MICOS complex was recently reported to be a central player determining cristae structure and formation of crista junctions. Here we investigated the functional role of MIC26, a lipoprotein formerly termed APOO. Its levels are increased in diabetic heart tissue and in blood plasma of patients suffering from acute coronary syndrome. We demonstrate that human MIC26 exists in three distinct forms: (1) a glycosylated and secreted 55kDa protein, (2) an ER/Golgi-resident form thereof, and (3) a non-glycosylated 22kDa mitochondrial protein. The latter isoform spans the mitochondrial inner membrane and physically interacts with several MICOS complex subunits such as MIC60, MIC27, and MIC10. We further demonstrate that MIC26 and MIC27, a homologous protein formerly termed APOOL, regulate their levels in an antagonistic manner. Both proteins are positively correlated with the levels of MIC10 as well as tafazzin, an enzyme required for cardiolipin remodeling. Overexpression of MIC26 induced fragmentation of mitochondria, promoted ROS formation and resulted in impaired mitochondrial respiration. Downregulation of MIC26 induced a decrease in mitochondrial oxygen consumption, whereas mitochondrial network morphology and ROS levels remained unaffected. MIC26 depletion led to alterations in mitochondrial ultrastructure and caused a significant reduction in the number of crista junctions. In summary, we show that the human apolipoprotein MIC26 is a bona fide subunit of the MICOS complex and that MIC26 is linked to cardiolipin metabolism and promotes crista junction formation.

We developed a rodent crista slice to investigate regional variations in electrophysiological properties of vestibular afferent terminals. Thin transverse slices of the gerbil crista ampullaris were made and electrical properties of calyx terminals in central zones (CZ) and peripheral zones (PZ) compared with whole cell patch clamp. Spontaneous action potential firing was observed in 25% of current-clamp recordings and was either regular or irregular in both zones. Firing was abolished when extracellular choline replaced Na+ but persisted when hair cell mechanotransduction channels or calyx AMPA receptors were blocked. This suggests that ion channels intrinsic to the calyx can generate spontaneous firing. In response to depolarizing voltage steps, outward K+ currents were observed at potentials above −60 mV. K+ currents in PZ calyces showed significantly more inactivation than currents in CZ calyces. Underlying K+ channel populations contributing to these differences were investigated. The KCNQ channel blocker XE991 dihydrochloride blocked a slowly activating, sustained outward current in both PZ and CZ calyces, indicating the presence of KCNQ channels. Mean reduction was greatest in PZ calyces. XE991 also reduced action potential firing frequency in CZ and PZ calyces and broadened mean action potential width. The K+ channel blocker 4-aminopyridine (10–50 μM) blocked rapidly activating, moderately inactivating currents that were more prevalent in PZ calyces. α-Dendrotoxin, a selective blocker of KV1 channels, reduced outward currents in CZ calyces but not in PZ calyces. Regional variations in K+ conductances may contribute to different firing responses in calyx afferents.

The differentiation of stem cells is a tightly regulated process essential for animal development and tissue homeostasis. Through this process, attainment of new identity and function is achieved by marked changes in cellular properties. Intrinsic cellular mechanisms governing stem cell differentiation remain largely unknown, in part because systematic forward genetic approaches to the problem have not been widely used1,2. Analysing genes required for germline stem cell differentiation in the Drosophila ovary, we find that the mitochondrial ATP synthase plays a critical role in this process. Unexpectedly, the ATP synthesizing function of this complex was not necessary for differentiation, as knockdown of other members of the oxidative phosphorylation system did not disrupt the process. Instead, the ATP synthase acted to promote the maturation of mitochondrial cristae during differentiation through dimerization and specific upregulation of the ATP synthase complex. Taken together, our results suggest that ATP synthase-dependent crista maturation is a key developmental process required for differentiation independent of oxidative phosphorylation.