Globular Cells Research Articles

Anatomy and histology of the metapleural gland in the giant tropical ant Paraponera clavata (Fabricius, 1775) (Formicidae: Paraponerinae).

Ants are social insects with about 85 exocrine glands with different morphologies. The metapleural gland is exclusive to ants and its compounds have antimicrobial action, indicating a function in the defense against pathogens. Within ants, Paraponera clavata is the only living species representative of Paraponerinae. This study describes the anatomy and histology of the metapleural gland in workers of P. clavata. The metapleural gland is formed by secretory units that release secretion in a reservoir through collecting canaliculi. Secretory units are bicellular with a well-developed globular secretory cell, with 145 µm in diameter, and a duct cell forming the collecting canaliculi characterizing a class III gland. The metapleural gland is pointed out as a synapomorphy of ants. Paraponerinae constitutes one of the six subfamilies of the basal and paraphyletic Poneroide clade and knowledge of the metapleural gland morphology contribute to the understanding of evolutionary history of the basal ants' groups.

Observation of Chronic Graft-Versus-Host Disease Mouse Model Cornea with In Vivo Confocal Microscopy.

Graft-versus-host disease (GVHD) is a major complication after hematopoietic stem cell transplantation (HSCT), and ocular GVHD can cause severe dry eye disease that can lead to visual impairment. Epithelial damage, vascular invasion, corneal fibrosis, and corneal perforation may occur in severe cases. It is generally accepted that inflammatory cells such as dendritic cells and T cells contribute to this pathological condition. However, it is still unknown what pathological condition occurs on the ocular surface after HSCT, and when. We therefore observed the dynamics of inflammatory cells in the cornea of chronic GVHD (cGVHD) model mice from 1 to 4 weeks after bone marrow transplantation (BMT) by in vivo confocal microscopy (IVCM) and considered the relationship with the pathophysiology of ocular GVHD (tear volume, corneal epithelial damage). In the allogeneic group, neovascularization occurred in all eyes at 1 week after BMT, although almost all vessels disappeared at 2 weeks after BMT. In addition, we revealed that infiltration of globular cells, and tortuosity and branching of nerves in the cornea occurred in both cGVHD mice and human cGVHD patients. Thus, we consider that cGVHD mouse model study by IVCM reproduces the state of ocular GVHD and may contribute to elucidating the pathological mechanism for ocular GVHD.

In-vitro Recordings of Neural Magnetic Activity From the Auditory Brainstem Using Color Centers in Diamond: A Simulation Study.

Magnetometry based on nitrogen-vacancy (NV) centers in diamond is a novel technique capable of measuring magnetic fields with high sensitivity and high spatial resolution. With the further advancements of these sensors, they may open up novel approaches for the 2D imaging of neural signals in vitro. In the present study, we investigate the feasibility of NV-based imaging by numerically simulating the magnetic signal from the auditory pathway of a rodent brainstem slice (ventral cochlear nucleus, VCN, to the medial trapezoid body, MNTB) as stimulated by both electric and optic stimulation. The resulting signal from these two stimulation methods are evaluated and compared. A realistic pathway model was created based on published data of the neural morphologies and channel dynamics of the globular bushy cells in the VCN and their axonal projections to the principal cells in the MNTB. The pathway dynamics in response to optic and electric stimulation and the emitted magnetic fields were estimated using the cable equation. For simulating the optic stimulation, the light distribution in brain tissue was numerically estimated and used to model the optogenetic neural excitation based on a four state channelrhodopsin-2 (ChR2) model. The corresponding heating was also estimated, using the bio-heat equation and was found to be low (<2°C) even at excessively strong optic signals. A peak magnetic field strength of ∼0.5 and ∼0.1 nT was calculated from the auditory brainstem pathway after electrical and optical stimulation, respectively. By increasing the stimulating light intensity four-fold (far exceeding commonly used intensities) the peak magnetic signal strength only increased to 0.2 nT. Thus, while optogenetic stimulation would be favorable to avoid artefacts in the recordings, electric stimulation achieves higher peak fields. The present simulation study predicts that high-resolution magnetic imaging of the action potentials traveling along the auditory brainstem pathway will only be possible for next generation NV sensors. However, the existing sensors already have sufficient sensitivity to support the magnetic sensing of cumulated neural signals sampled from larger parts of the pathway, which might be a promising intermediate step toward further maturing this novel technology.

Gastruloid Development Competence Discriminates Different States of Pluripotency.

SummaryFloating spheroidal aggregates of mouse embryonic stem cells can develop into polarized/elongated organoids, namely gastruloids. We set up a high-performing assay to measure gastruloid formation efficiency (GFE), and found that GFE decreases as pluripotency progresses from naive (GFE ≥ 95%) to primed (GFE = 0) state. Specifically, we show that primed EpiSCs fail to generate proper cell aggregates, while early-primed EpiLCs aggregate but eventually fail to develop into elongated gastruloids. Moreover, we characterized proline-induced cells (PiCs), a LIF-dependent reversible early-primed state of pluripotency, and show that PiCs are able to generate gastruloids (GFE ∼ 50%) and are also competent to differentiate into primordial germ cell-like cells. Thus, we propose the GFE assay as a valuable functional tool to discriminate different states of the pluripotency continuum.

Endocannabinoids regulate cerebellar GABAergic transmission in a synapse type‐dependent manner

Endocannabinoids (eCBs) act as ubiquitous modulators of synaptic transmission via the activation of cannabinoid receptors (CBRs). Cerebellar Purkinje cells (PCs) make strong inhibitory synaptic contacts not only with neurons in the deep cerebellar nuclei (DCN) but also with Lugaro cells and globular cells, whose cell bodies are located underneath the PC layer. However, little is known about the modulatory actions of eCBs on GABA release from PC axon terminals. Here, we examined the effects of eCBs on the GABAergic transmission at PC-globular cell synapses and PC-large DCN neuron synapses electrophysiologically using mouse cerebellar slices. We showed that the types 1 and 2 CBR agonist WIN55212 did not affect either spontaneous or miniature inhibitory postsynaptic currents (IPSCs) in globular cells under control conditions and in a state of enhanced synaptic activity. By contrast, another Gi/o protein-coupled receptor agonist, baclofen, significantly reduced the miniature IPSC frequency in globular cells. WIN55212 had no effects on IPSCs in large DCN neurons. A type 2 CBR agonist, HU308, also had no effects on IPSCs in either globular cells or large DCN neurons. Moreover, the PCs' target neurons did not elicit depolarization-induced suppression of inhibition. These results suggest the lack of a functional role of CBRs at PCs' axon terminals. This is in sharp contrast to the fact that PCs receive abundant excitatory and inhibitory inputs that are under eCB-mediated presynaptic inhibitory modulation. The actions of eCBs are selective to distinct synapses and possibly contribute to information processes and rigorous signal transmission in the cerebellum.

Computational modeling of the auditory brainstem response to continuous speech

Objective. The auditory brainstem response can be recorded non-invasively from scalp electrodes and serves as an important clinical measure of hearing function. We have recently shown how the brainstem response at the fundamental frequency of continuous, non-repetitive speech can be measured, and have used this measure to demonstrate that the response is modulated by selective attention. However, different parts of the speech signal as well as several parts of the brainstem contribute to this response. Here we employ a computational model of the brainstem to elucidate the influence of these different factors. Approach. We developed a computational model of the auditory brainstem by combining a model of the middle and inner ear with a model of globular bushy cells in the cochlear nuclei and with a phenomenological model of the inferior colliculus. We then employed the model to investigate the neural response to continuous speech at different stages in the brainstem, following the methodology developed recently by ourselves for detecting the brainstem response to running speech from scalp recordings. We compared the simulations with recordings from healthy volunteers. Main results. We found that the auditory-nerve fibers, the cochlear nuclei and the inferior colliculus all contributed to the speech-evoked brainstem response, although the dominant contribution came from the inferior colliculus. The delay of the response corresponded to that observed in experiments. We further found that a broad range of harmonics of the fundamental frequency, up to about 8 kHz, contributed to the brainstem response. The response declined with increasing fundamental frequency, although the signal-to-noise ratio was largely unaffected. Significance. Our results suggest that the scalp-recorded brainstem response at the fundamental frequency of speech originates predominantly in the inferior colliculus. They further show that the response is shaped by a large number of higher harmonics of the fundamental frequency, reflecting highly nonlinear processing in the auditory periphery and illustrating the complexity of the response.

Structure and dynamics of hydrodynamically interacting finite-size Brownian particles in a spherical cavity: Spheres and cylinders.

The structure and dynamics of confined suspensions of particles of arbitrary shape are of interest in multiple disciplines from biology to engineering. Theoretical studies are often limited by the complexity of long-range particle-particle and particle-wall forces, including many-body fluctuating hydrodynamic interactions. Here, we report a computational study on the diffusion of spherical and cylindrical particles confined in a spherical cavity. We rely on an immersed-boundary general geometry Ewald-like method to capture lubrication and long-range hydrodynamics and include appropriate non-slip conditions at the confining walls. A Chebyshev polynomial approximation is used to satisfy the fluctuation-dissipation theorem for the Brownian suspension. We explore how lubrication, long-range hydrodynamics, particle volume fraction, and shape affect the equilibrium structure and the diffusion of the particles. It is found that once the particle volume fraction is greater than 10%, the particles start to form layered aggregates that greatly influence particle dynamics. Hydrodynamic interactions strongly influence the particle diffusion by inducing spatially dependent short-time diffusion coefficients, stronger wall effects on the particle diffusion toward the walls, and a sub-diffusive regime-caused by crowding-in the long-time particle mobility. The level of asymmetry of the cylindrical particles considered here is enough to induce an orientational order in the layered structure, decreasing the diffusion rate and facilitating a transition to the crowded mobility regime at low particle concentrations. Our results offer fundamental insights into the diffusion and distribution of globular and fibrillar proteins inside cells.

Morphology and chemical composition of the Koschewnikow gland of the honey bee Apis mellifera (Hymenoptera: Apidae) workers engaged in different tasks

The Koschewnikow gland is associated with the sting apparatus in the honey bee Apis mellifera Linnaeus, 1758, as a potential source of compounds used as an alarm pheromone. This study describes the histology, morphometry and ultrastructure, and chemical composition of the Koschewnikow gland in workers performing different tasks in the A. mellifera colony. The Koschewnikow gland shows well-developed globular secretory cells of class III with those of the nurse and foragers larger and in greater numbers than those of guard bees. At the ultrastructural level, the secretory cells of all worker bees are rich in mitochondria and smooth endoplasmic reticulum. Mitochondria had different ultrastructure among workers with membrane electron-dense granules in nurse and the electron-lucent halo surrounding the organelle in guard bees. Mitochondria in forager workers show degenerative features such as the disappearance of cristae, electron-lucent matrix, and accumulation of electron-dense aggregates. The chemical analyses of the Koschewnikow gland extract showed that of 23 compounds, 13 had different abundances between workers and of these, 10 were found in higher abundance in guard workers. These results suggest that the Koschewnikow gland has a different structure and chemical composition in honey bee workers engaged in different tasks.

Neuronal population model of globular bushy cells covering unit-to-unit variability.

Computations of acoustic information along the central auditory pathways start in the cochlear nucleus. Bushy cells in the anteroventral cochlear nucleus, which innervate monaural and binaural stations in the superior olivary complex, process and transfer temporal cues relevant for sound localization. These cells are categorized into two groups: spherical and globular bushy cells (SBCs/GBCs). Spontaneous rates of GBCs innervated by multiple auditory nerve (AN) fibers are generally lower than those of SBCs that receive a small number of large AN synapses. In response to low-frequency tonal stimulation, both types of bushy cells show improved phase-locking and entrainment compared to AN fibers. When driven by high-frequency tones, GBCs show primary-like-with-notch or onset-L peristimulus time histograms and relatively irregular spiking. However, previous in vivo physiological studies of bushy cells also found considerable unit-to-unit variability in these response patterns. Here we present a population of models that can simulate the observed variation in GBCs. We used a simple coincidence detection model with an adaptive threshold and systematically varied its six parameters. Out of 567000 parameter combinations tested, 7520 primary-like-with-notch models and 4094 onset-L models were selected that satisfied a set of physiological criteria for a GBC unit. Analyses of the model parameters and output measures revealed that the parameters of the accepted model population are weakly correlated with each other to retain major GBC properties, and that the output spiking patterns of the model are affected by a combination of multiple parameters. Simulations of frequency-dependent temporal properties of the model GBCs showed a reasonable fit to empirical data, supporting the validity of our population modeling. The computational simplicity and efficiency of the model structure makes our approach suitable for future large-scale simulations of binaural information processing that may involve thousands of GBC units.

Isolation of cancer stem cells and the establishment of a H 2O 2-resistant cancer stem cell model

To isolate cancer stem cells (CST) from human breast cancer cell line (MCF-7) and study their sensitivity toward oxidative stress. MCF-7 cells were cultured in serum-free suspension culture medium to identify cells forming the sphere phenotype. The morphological changes of MCF-7 cells were observed by inverted phase contrast microscope (compared with MCF-7 cells cultured in serum-free suspension culture medium). The expression of CST marker CD133 was detected by immunocytochemical staining in CST cell spheres (experimental group) with a diameter of 100 μm and MCF-7 cells (control group) with a fusion degree of 70%. The positive rate of CD133 was detected by flow cytometry in the third generation of tumor cells with diameter of 150 μm. The second generation of tumor globular cells (experimental group) with diameter of 150 μm and corresponding MCF-7 cells (control group) were taken to be damaged by 50 mol/L H 2O 2 for 120 minutes. The expression of DNA damage marker histone H2AX phosphorylation (γH2AX) was detected by immunocytochemical staining. Inverted phase contrast microscopy showed that MCF-7 cells grew initially in a single-cell adherent state, then aggregated and grew in serum-free suspension culture medium, and finally formed CST cell spheres, while the control MCF-7 cells cultured in MCF-7 cell culture medium grew extensively and could not grow in suspension. Fluorescence microscopy showed that the expression of CD133 in MCF-7 cells of control group was negative, while that in experimental group was positive. Flow cytometry showed that CD133 was positive in CST cells, and the positive rate was 92%. Inverted fluorescence microscopy showed that the expression of γH2AX in CST tumor spheres of experimental group was significantly lower than that in MCF-7 cells of control group after 120 minutes of H 2O 2 injury. Serum-free suspension culture medium can produce globular CST cells from MCF-7 tumor cell line, which have strong antioxidant damage.

Rapid and Efficient Conversion of Human Fibroblasts into Functional Neurons by Small Molecules.

SummaryRecent studies have demonstrated that human astrocytes and fibroblasts can be directly converted into functional neurons by small molecules. However, fibroblasts, as a potentially better cell resource for transplantation, are not as easy to reprogram as astrocytes regarding their fate to neurons, and chemically induced neurons (iNs) with low efficiency from fibroblasts resulted in limited application for the treatment of neurological disorders, including depression. Here, we report that human fibroblasts can be efficiently and directly reprogrammed into glutamatergic neuron-like cells by serially exposing cells to a combination of small molecules. These iNs displayed neuronal transcriptional networks, and also exhibited mature firing patterns and formed functional synapses. Importantly, iNs could integrate into local circuits after transplantation into postnatal mouse brain. Our study provides a rapid and efficient transgene-free approach for chemically generating neuron-like cells from human fibroblasts. Furthermore, our approach offers strategies for disease modeling and drug discovery in central nervous system disorders.

Application of spherical substrate to observe bacterial motility machineries by Quick-Freeze-Replica Electron Microscopy

3-D Structural information is essential to elucidate the molecular mechanisms of various biological machineries. Quick-Freeze Deep-Etch-Replica Electron Microscopy is a unique technique to give very high-contrast surface profiles of extra- and intra-cellular apparatuses that bear numerous cellular functions. Though the global architecture of those machineries is primarily required to understand their functional features, it is difficult or even impossible to depict side- or highly-oblique views of the same targets by usual goniometry, inasmuch as the objects (e.g. motile microorganisms) are placed on conventional flat substrates. We introduced silica-beads as an alternative substrate to solve such crucial issue. Elongated Flavobacterium and globular Mycoplasmas cells glided regularly along the bead’s surface, similarly to those on a flat substrate. Quick-freeze replicas of those cells attached to the beads showed various views; side-, oblique- and frontal-views, enabling us to study not only global but potentially more detailed morphology of complicated architecture. Adhesion of the targets to the convex surface could give surplus merits to visualizing intriguing molecular assemblies within the cells, which is relevant to a variety of motility machinery of microorganisms.

Histological comparison of in vitro and in vivo development of peripheral posterior capsule opacification in human donor tissue

In order to study the mechanisms involved in the development of posterior capsule opacification (PCO) we compared in vivo developed PCO with PCO formed in tissue culture with focus on the periphery of the lens capsule to evaluate lens regeneration potential. We studied three human tissue groups: Cultured lens capsules after mock cataract surgery (n = 6, 30 days), lens capsules from donors that had previously undergone cataract surgery (IOL capsules) (n = 12) and intact lenses (n = 6). All samples were stained with Vimentin, alpha Smooth Muscle Actin, Picro Sirius Red (for collagen) and Paired box protein (Pax6). We found that cultured capsules and less developed IOL capsules consisted mainly of monolayers of mesenchymal cells, while more developed IOL capsules, contained lens epithelial cells (LECs), globular cells and lens fiber cells. Many IOL capsule samples expressed collagen I and III in areas where cells were in contact with the IOL. Pax6 had a similar dispersed distribution in less developed IOL capsules and cultured capsules, while more developed IOL capsules and intact lenses, concentrated Pax6 in LECs at the equatorial lens bow. The similarities between cultured capsules and less developed IOL capsules indicate that our in vitro developed PCO is comparable to early in vivo developed PCO. The similar morphology of more developed IOL capsules and intact lenses seems to indicate an attempt at lens regeneration.

A Comparative Histomorphological and Histochemical Study of the Goblet Cells and Brunner's Glands in the Duodenum of Rabbits and Rats

The purpose of this study was to describe, comparison the histological structures, histochemical features and distribution of the goblet cells and Brunner's glands in the duodenum of the rabbits and rats. The study was carried out on Ten samples of duodenum of each of rabbits and rats, immediately after slaughtering. The specimens were divided into cranial, descending and ascending parts. Five specimens was taken from each part of duodenum of each of the rabbits and rats and fixed in 10% formalin for 24 hours and treating by routine histological technique. Sections were stained by Hematoxylin and Eosin, Picric acid stain and Alcian blue PH 2.5 stains. Mean length of crypts of Lieberkuhn and number of each crypts, goblet cells and alveoli of Brunner's glands in duodenum of rabbits were more than that in the duodenum of rats. The mean length and number of crypts of Lieberkuhn was increase toward the last parts of the duodenum in each of the rabbits and rats. Goblet cells were globular cells between the columnar cells in the epithelium lined each of the villi and crypts of Lieberkuhn in the tunica mucosa of the duodenum in rabbits and rats and take positive reaction each of PAS and Alcian blue pH 2.5 stains. The mean number of goblet cells in crypts of Lieberkuhn in duodenum each rabbits and rats were increased toward the jejunum. Brunner's glands were branched tubuloalveolar glands in the submucosa of each duodenal parts in rabbits, while absent in ascending part of rats duodenum, and these glands in rabbits duodenum were composed of two types of the cells (serous and mucous), while in rats duodenum only mucous cells, and stained with PAS stain only in duodenum of each of the rabbits and rats. The mean number of the alveoli of the Brunner's glands in the cranial part were more than that in other parts and decreased toward ascending part in rabbits duodenum, while absent in ascending part of rats duodenum.

Predicting the vascular adhesion of deformable drug carriers in narrow capillaries traversed by blood cells

In vascular targeted therapies, blood-borne carriers should realize sustained drug release from the luminal side towards the diseased tissue. In this context, such carriers are required to firmly adhere to the vessel walls for a sufficient period of time while resisting force perturbations induced by the blood flow and circulating cells. Here, a hybrid computational model, combining a Lattice Boltzmann (LBM) and Immersed Boundary Methods (IBM), is proposed for predicting the strength of adhesion of particles in narrow capillaries (7.5μm) traversed by blood cells. While flowing down the capillary, globular and biconcave deformable cells (7μm) encounter 2μm discoidal particles, adhering to the vessel walls. Particles present aspect ratios ranging from 0.25 to 1.0 and a mechanical stiffness varying from rigid (Ca = 0) to soft (Ca = 10−3). Cell–particle interactions are quantitatively predicted over time via three independent parameters: the cell membrane stretching δp; the cell-to-particle distance r, and the number of engaged ligand–receptor bonds NL. Under physiological flow conditions (Re = 10−2), rigid particles are detached and displaced away from the wall by blood cells. This is associated with a significant cell membrane stretching (up to 10%) and rapid breaking of molecular bonds (t umax/H <1). Differently, soft particles deform their shape as cells pass by, thus reducing force perturbations and extending the life of molecular bonds. Yet, only the thinnest deformable particles (2 ×0.5μm) firmly adhere to the walls under all tested configurations. These results suggest that low aspect ratio deformable particles can establish long-lived adhesive interactions with capillary walls, enabling de facto vascular targeted therapies.

First description of extrafloral nectaries in Opuntia robusta (Cactaceae): Anatomy and ultrastructure.

To our knowledge, there are no studies about the structure and ecological function of extrafloral nectaries (EFNs) in Opuntia robusta. This is the first description of EFNs in O. robusta, where young spines have an interesting structure and a secreting function, which are different from EFNs described in other Cactaceae species. We used light, scanning-electron, and transmission-electron microscopy to examine morphology, anatomy, and ultrastructure of the secretory spines in areoles in female and hermaphrodite individuals of O. robusta. Young cladodes develop areoles with modified and secretory spines as EFNs only active during the early growth phase. EFNs are non-vascularized structures, with no stomata, that consist of a basal meristematic tissue, a middle elongation region, and an apical secretory cone formed by large globular epidermal cells, containing nectar and medullar elongated cells. We observed the presence of Golgi apparatus, vesicles and plastids in the medullar and sup-epidermal cells of the spine. We propose that the nectar is stored in the globular cells at the apex of the spine and secreted by breaking through the globular cells or by pores. We recorded a more frequent presence of ants on younger cladode sprouts producing young secreting spines: this result is parallel with the predictions of Optimal Defense Hypothesis, which states that younger plant organs should be better defended than older ones because their loss produces a higher fitness impairment. Although Diaz-Castelazo’s hypothesis states that a more complex structure of EFNs correlates with their lower among-organs dispersion, comparing to less complex EFNs, non-vascularized structure of EFNs in O. robusta is not associated with their higher among-organs dispersion likened to O. stricta, which produces vascularized EFNs. We provide evidence that this characteristic is not a good taxonomic feature of Opuntia genus. Moreover, the comparison of EFNs of O. robusta and O. stricta suggests that the hypothesis of Diaz-Castelazo should be revised: it is rather a rule but not a law.

Effect of ancymidol on cell wall metabolism in growing maize cells.

Ancymidol inhibits the incorporation of cellulose into cell walls of maize cell cultures in a gibberellin-independent manner, impairing cell growth; the reduction in the cellulose content is compensated with xylans. Ancymidol is a plant growth retardant which impairs gibberellin biosynthesis. It has been reported to inhibit cellulose synthesis by tobacco cells, based on its cell-malforming effects. To ascertain the putative role of ancymidol as a cellulose biosynthesis inhibitor, we conducted a biochemical study of its effect on cell growth and cell wall metabolism in maize cultured cells. Ancymidol concentrations ≤500µM progressively reduced cell growth and induced globular cell shape without affecting cell viability. However, cell growth and viability were strongly reduced by ancymidol concentrations≥1.5mM. The I50 value for the effect of ancymidol on FW gain was 658µM. A reversal of the inhibitory effects on cell growth was observed when 500µM ancymidol-treated cultures were supplemented with 100µM GA3. Ancymidol impaired the accumulation of cellulose in cell walls, as monitored by FTIR spectroscopy. Cells treated with 500µM ancymidol showed a ~60% reduction in cellulose content, with no further change as the ancymidol concentration increased. Cellulose content was partially restored by 100µM GA3. Radiolabeling experiments confirmed that ancymidol reduced the incorporation of [14C]glucose into α-cellulose and this reduction was not reverted by the simultaneous application of GA3. RT-PCR analysis indicated that the cellulose biosynthesis inhibition caused by ancymidol is not related to a downregulation of ZmCesA gene expression. Additionally, ancymidol treatment increased the incorporation of [3H]arabinose into a hemicellulose-enriched fraction, and up-regulated ZmIRX9 and ZmIRX10L gene expression, indicating an enhancement in the biosynthesis of arabinoxylans as a compensatory response to cellulose reduction.

Histological evidence of a glandular tubercle at the abdominal tergite in three species of katydids (Orthoptera: Tettigoniidae)

Three species of Hyperophora Brunner von Wattenwyl have a tubercle on the second abdominal tergite with different external appearance among species. We predicted that the tubercles are a glandular structure for their morphology and anatomical position. In H. major Brunner von Wattenwyl and H. brasiliensis Brunner von Wattenwyl the tubercle is low and broad, whereas in H. minor Brunner von Wattenwyl it is high. In all species the tubercle does not have an external opening, but it has many chemosensory sensilla, evidenced by long setae with an internal cavity with a membrane-like content. The cuticular surface of the tubercle has many pores with ∼1–2μm in diameter scattered among the sensilla. On the inner surface of the tubercular region occur epidermal gland cells, evidenced by a mass of large globular cells with an irregular and well-developed nucleus with predominance of decondensed chromatin and a large nucleolus. The cytoplasm of these cells is rich in small clear vacuoles, and has an end apparatus, a feature of epidermal gland cells of class III. Each gland cell has a conducting canal that opens into a cuticular pore. These results support the hypothesis that the tubercles are indeed of glandular nature.

ER-PM Contacts Restrict Exocytic Sites for Polarized Morphogenesis

Spatial control of exocytosis underlies polarized cell morphogenesis. In rod-shaped fission yeast, exocytic vesicles are conveyed along the actin cytoskeleton by myosin V motors toward growing cell ends [1,2], the major sites for exocytosis. However, actomyosin-based vesicle delivery is dispensable for polarized secretion and cylindrical cell shape of fission yeast [3]. Thus, additional mechanisms should function in the spatial confinement of exocytosis. Here we report a novel role of endoplasmic reticulum (ER)-plasma membrane (PM) contacts in restricting exocytic sites for polarized fission yeast morphogenesis. We show that fission yeast cells deficient in both ER-PM contacts and actomyosin-based secretory vesicle transport display aberrant globular cell shape due to delocalized exocytosis. By artificially manipulating the strength and extent of ER-PM contacts in wild-type and mutant cells that exhibit induced ectopic exocytosis, we demonstrate that exocytosis and ER-PM contact formation are spatially incompatible. Furthermore, extensive ER-PM junctions at the non-growing lateral cell cortex prevent the PM from exocytic vesicle tethering and hence attenuate growth potential at cell sides. We thus propose that ER-PM contacts function as a new morphogenetic module by limiting exocytosis to growing cell tips in fission yeast. A similar mechanism could apply to other cell types with prominent ER-PM contacts.

SK Channels Regulate Resting Properties and Signaling Reliability of a Developing Fast-Spiking Neuron.

Reliable and precise signal transmission is essential in circuits of the auditory brainstem to encode timing with submillisecond accuracy. Globular bushy cells reliably and faithfully transfer spike signals to the principal neurons of the medial nucleus of the trapezoid body (MNTB) through the giant glutamatergic synapse, the calyx of Held. Thus, the MNTB works as a relay nucleus that preserves the temporal pattern of firing at high frequency. Using whole-cell patch-clamp recordings, we observed a K+ conductance mediated by small-conductance calcium-activated potassium (SK) channels in the MNTB neurons from rats of either sex. SK channels were activated by intracellular Ca2+ sparks and mediated spontaneous transient outward currents in developing MNTB neurons. SK channels were also activated by Ca2+ influx through voltage-gated Ca2+ channels and synaptically activated NMDA receptors. Blocking SK channels with apamin depolarized the resting membrane potential, reduced resting conductance, and affected the responsiveness of MNTB neurons to signal inputs. Moreover, SK channels were activated by action potentials and affected the spike afterhyperpolarization. Blocking SK channels disrupted the one-to-one signal transmission from presynaptic calyces to postsynaptic MNTB neurons and induced extra postsynaptic action potentials in response to presynaptic firing. These data reveal that SK channels play crucial roles in regulating the resting properties and maintaining reliable signal transmission of MNTB neurons.SIGNIFICANCE STATEMENT Reliable and precise signal transmission is required in auditory brainstem circuits to localize the sound source. The calyx of Held synapse in the mammalian medial nucleus of the trapezoid body (MNTB) plays an important role in sound localization. We investigated the potassium channels that shape the reliability of signal transfer across the calyceal synapse and observed a potassium conductance mediated by small-conductance calcium-activated potassium (SK) channels in rat MNTB principal neurons. We found that SK channels are tonically activated and contribute to the resting membrane properties of MNTB neurons. Interestingly, SK channels are transiently activated by calcium sparks and calcium influx during action potentials and control the one-to-one signal transmission from presynaptic calyces to postsynaptic MNTB neurons.