Mks6 mutations reveal tissue- and cell type-specific roles for the cilia transition zone.

The transition zone (TZ) is a domain at the base of the cilium that is involved in maintaining ciliary compartment-specific sensory and signaling activity by regulating cilia protein composition. Mutations in TZ proteins result in cilia dysfunction, often causing pleiotropic effects observed in a group of human diseases classified as ciliopathies. The purpose of this study is to describe the importance of the TZ component Meckel-Grüber syndrome 6 ( Mks6) in several organ systems and tissues regarding ciliogenesis and cilia maintenance using congenital and conditional mutant mouse models.

Mutation of Growth Arrest Specific 8 Reveals a Role in Motile Cilia Function and Human Disease.

Ciliopathies are genetic disorders arising from dysfunction of microtubule-based cellular appendages called cilia. Different cilia types possess distinct stereotypic microtubule doublet arrangements with non-motile or 'primary' cilia having a 9+0 and motile cilia have a 9+2 array of microtubule doublets. Primary cilia are critical sensory and signaling centers needed for normal mammalian development.

A Screen for Modifiers of Cilia Phenotypes Reveals Novel MKS Alleles and Uncovers a Specific Genetic Interaction between osm-3 and nphp-4.

Nephronophthisis (NPHP) is a ciliopathy in which genetic modifiers may underlie the variable penetrance of clinical features. To identify modifiers, a screen was conducted on C. elegans nphp-4(tm925) mutants.

KIAA0556 is a novel ciliary basal body component mutated in Joubert syndrome.

Background: Joubert syndrome (JBTS) and related disorders are defined by cerebellar malformation (molar tooth sign), together with neurological symptoms of variable expressivity. The ciliary basis of Joubert syndrome related disorders frequently extends the phenotype to tissues such as the eye, kidney, skeleton and craniofacial structures.

Results: Using autozygome and exome analyses, we identified a null mutation in KIAA0556 in a multiplex consanguineous family with hallmark features of mild Joubert syndrome.

Hippocampal and cortical primary cilia are required for aversive memory in mice.

It has been known for decades that neurons throughout the brain possess solitary, immotile, microtubule based appendages called primary cilia. Only recently have studies tried to address the functions of these cilia and our current understanding remains poor. To determine if neuronal cilia have a role in behavior we specifically disrupted ciliogenesis in the cortex and hippocampus of mice through conditional deletion of the Intraflagellar Transport 88 (Ift88) gene.

An inducible CiliaGFP mouse model for in vivo visualization and analysis of cilia in live tissue.

Background: Cilia are found on nearly every cell type in the mammalian body, and have been historically classified as either motile or immotile. Motile cilia are important for fluid and cellular movement; however, the roles of non-motile or primary cilia in most tissues remain unknown. Several genetic syndromes, called the ciliopathies, are associated with defects in cilia structure or function and have a wide range of clinical presentations.

Leptin resistance is a secondary consequence of the obesity in ciliopathy mutant mice.

Although primary cilia are well established as important sensory and signaling structures, their function in most tissues remains unknown. Obesity is a feature associated with some syndromes of cilia dysfunction, such as Bardet-Biedl syndrome (BBS) and Alström syndrome, as well as in several cilia mutant mouse models. Recent data indicate that obesity in BBS mutant mice is due to defects in leptin receptor trafficking and leptin resistance.

Proximal tubule proliferation is insufficient to induce rapid cyst formation after cilia disruption.

Disrupting the function of cilia in mouse kidneys results in rapid or slow progression of cystic disease depending on whether the animals are juveniles or adults, respectively. Renal injury can also markedly accelerate the renal cyst formation that occurs after disruption of cilia in adult mice. Rates of cell proliferation are markedly higher in juvenile than adult kidneys and increase after renal injury, suggesting that cell proliferation may enhance the development of cysts.

Method for the determination of trajectory angles of directional secretory vesicles in cultured astrocytes.

Astrocytes provide a principal pathway for glutamate uptake in the mammalian brain, a task accomplished by the powerful action of excitatory amino acid transporters (EAAT) 1 and 2. These transporters are synthesized within the endoplasmic reticulum and are then trafficked to the plasma membrane. The characteristics of their intracellular traffic within astrocytes have not been investigated.

Microtubule modifications and stability are altered by cilia perturbation and in cystic kidney disease.

Disruption of the primary cilium is associated with a growing number of human diseases collectively termed ciliopathies. Ciliopathies present with a broad range of clinical features consistent with the near ubiquitous nature of the organelle and its role in diverse signaling pathways throughout development and adult homeostasis. The clinical features associated with cilia dysfunction can include such phenotypes as polycystic kidneys, skeletal abnormalities, blindness, anosmia, and obesity.

Temporal characteristics of vesicular fusion in astrocytes: examination of synaptobrevin 2-laden vesicles at single vesicle resolution.

Astrocytes can release various gliotransmitters in response to stimuli that cause increases in intracellular Ca(2+) levels; this secretion occurs via a regulated exocytosis pathway. Indeed, astrocytes express protein components of the vesicular secretory apparatus. However, the detailed temporal characteristics of vesicular fusions in astrocytes are not well understood.

Role of epidermal primary cilia in the homeostasis of skin and hair follicles.

Skin and hair follicle morphogenesis and homeostasis require the integration of multiple signaling pathways, including Hedgehog (Hh) and Wingless (Wnt), and oriented cell divisions, all of which have been associated with primary cilia. Although studies have shown that disrupting dermal cilia causes follicular arrest and attenuated Hh signaling, little is known about the role of epidermal cilia. Here, epidermal cilia function was analyzed using conditional alleles of the ciliogenic genes Ift88 and Kif3a.

Carbon nanotubes in neuroscience.

Carbon nanotubes have electrical, mechanical and chemical properties that make them one of the most promising materials for applications in neuroscience. Single-walled and multi-walled carbon nanotubes have been increasingly used as scaffolds for neuronal growth and more recently for neural stem cell growth and differentiation. They are also used in interfaces with neurons, where they can detect neuronal electrical activity and also deliver electrical stimulation to these cells.

Dynamic imaging of cannabinoid receptor 1 vesicular trafficking in cultured astrocytes.

Astrocytes possess GPCRs (G-protein-coupled receptors) for neuroactive substances and can respond via these receptors to signals originating from neurons as well as astrocytes. Like many transmembrane proteins, GPCRs exist in a dynamic equilibrium between receptors expressed at the plasma membrane and those present within intracellular trafficking compartments. The characteristics of GPCR trafficking within astrocytes have not been investigated.

Conductive single-walled carbon nanotube substrates modulate neuronal growth.

We used conductive nanotube films as substrates with which we could systematically vary the conductance to see how this property affects neuronal growth. Here we show that nanotube substrates in a narrow range of conductivity promote the outgrowth of neurites with a decrease in the number of growth cones as well as an increase in cell body area, while at higher conductance these effects disappear.

Micropit: A New Cell Culturing Approach for Characterization of Solitary Astrocytes and Small Networks of these Glial Cells.

Astrocytes play an important role in cell-cell signaling in the mammalian central nervous system. The ability of astrocytes to communicate with surrounding cells through gap-junctional coupling or signaling via the release of transmitters makes characterization of these cells difficult in vitro and even more so in vivo. To simplify the complexity of common in vitro systems, introduced by intercellular communication between astrocytes, we developed a novel cell culturing method, in which purified rat visual cortical astrocytes were grown in spatially defined cell-adhesion wells which we termed micropits.

Water soluble single-walled carbon nanotubes inhibit stimulated endocytosis in neurons.

We report the use of chemically functionalized water soluble single-walled carbon nanotube (SWNT) graft copolymers to inhibit endocytosis. The graft copolymers were prepared by the functionalization of SWNTs with polyethylene glycol. When added to the culturing medium, these functionalized water soluble SWNTs were able to increase the length of various neuronal processes, neurites, as previously reported.

Ca2+ entry through TRPC1 channels contributes to intracellular Ca2+ dynamics and consequent glutamate release from rat astrocytes.

Astrocytes can respond to a variety of stimuli by elevating their cytoplasmic Ca2+ concentration and can in turn release glutamate to signal adjacent neurons. The majority of this Ca2+ is derived from internal stores while a portion also comes from outside of the cell. Astrocytes use Ca2+ entry through store-operated Ca2+ channels to refill their internal stores.

Vesicular release of glutamate mediates bidirectional signaling between astrocytes and neurons.

The major excitatory neurotransmitter in the CNS, glutamate, can be released exocytotically by neurons and astrocytes. Glutamate released from neurons can affect adjacent astrocytes by changing their intracellular Ca(2+) dynamics and, vice versa, glutamate released from astrocytes can cause a variety of responses in neurons such as: an elevation of [Ca(2+)](i), a slow inward current, an increase of excitability, modulation of synaptic transmission, synchronization of synaptic events, or some combination of these. This astrocyte-neuron signaling pathway might be a widespread phenomenon throughout the brain with astrocytes possessing the means to be active participants in many functions of the CNS.

Mechanisms of glutamate release from astrocytes.

Astrocytes can release the excitatory transmitter glutamate which is capable of modulating activity in nearby neurons. This astrocytic glutamate release can occur through six known mechanisms: (i) reversal of uptake by glutamate transporters (ii) anion channel opening induced by cell swelling, (iii) Ca2+-dependent exocytosis, (iv) glutamate exchange via the cystine-glutamate antiporter, (v) release through ionotropic purinergic receptors and (vi) functional unpaired connexons, "hemichannels", on the cell surface. Although these various pathways have been defined, it is not clear how often and to what extent astrocytes employ different mechanisms.

Applications of carbon nanotubes in neurobiology.

Background: Carbon nanotubes are one of the most promising materials for the electronics, computer and aerospace industries. There are numerous properties of carbon nanotubes that make them attractive for applications in neurobiology: small size, flexibility, strength, inertness, electrical conductivity and ease of modification with biological compounds.

Objective/methods: Here, we discuss the current applications of carbon nanotubes in neuroscience.

Vesicular transmitter release from astrocytes.

Astrocytes can release a variety of transmitters, including glutamate and ATP, in response to stimuli that induce increases in intracellular Ca(2+) levels. This release occurs via a regulated, exocytotic pathway. As evidence of this, astrocytes express protein components of the vesicular secretory apparatus, including synaptobrevin 2, syntaxin, and SNAP-23.

Chemically functionalized water soluble single-walled carbon nanotubes modulate neurite outgrowth.

We report the use of chemically-functionalized water soluble single-walled carbon nanotube (SWNT) graft copolymers for modulation of outgrowth of neuronal processes. The graft copolymers were prepared by the functionalization of SWNTs with poly-m-aminobenzene sulphonic acid and polyethylene glycol. When added to the culturing medium, these functionalized water soluble SWNTs were able to increase the length of various neuronal processes.

Applications of Carbon Nanotubes in Biotechnology and Biomedicine.

Due to their electrical, chemical, mechanical and thermal properties, carbon nanotubes are one of the most promising materials for the electronics, computer and aerospace industries. Here, we discuss their properties in the context of future applications in biotechnology and biomedicine. The purification and chemical modification of carbon nanotubes with organic, polymeric and biological molecules are discussed.