William D Phillips

The University of Sydney · Bosch Institute

Side population (SP) cells are a small subpopulation of cells found in many mammalian tissues and organs, identified by their capacity to efflux Hoechst 33342 dye. They are enriched for stem/progenitor cell activity. SP cells isolated from the adult mouse lung can be separated into a CD45+ subset (bone marrow-derived) and a CD45⁻ subset that can be...

Cannabinoids exert dynamic control over many physiological processes including memory formation, cognition and pain perception. In the central nervous system endocannabinoids mediate negative feedback of quantal transmitter release following postsynaptic depolarization. The influence of cannabinoids in the peripheral nervous system is less clear an...

Myasthenia gravis (MG) is an autoimmune disorder characterized by generalized muscle weakness due to autoantibodies causing dysfunction of the neuromuscular junction (NMJ). Steroids and other immunosuppressant’s provide effective treatment for MG. However, these treatments often cause severe side effects, and some patients are treatment resistant....

While the majority of myasthenia gravis patients express antibodies targeting the acetylcholine receptor, the second most common cohort instead displays autoantibodies against muscle-specific kinase (MuSK). MuSK is a transmembrane tyrosine kinase found in the postsynaptic membrane of the neuromuscular junction. During development, MuSK serves as a...

Background: The L25 mouse line was generated by random genomic insertion of a lens-specific transgene. Inbreeding of L25 hemizygotes revealed an unanticipated spastic phenotype in the hind limbs. Objective: The goals were to characterize the motor phenotype in the L25 mice and to map the transgene insert site within the mouse genome. Methods:...

Background: Side-population (SP) cells, identified by their capacity to efflux Hoechst dye, are highly enriched for stem/progenitor cell activity. They are found in many mammalian tissues, including mouse heart. Studies suggest that cardiac SP (CSP) cells can be divided into SCA1(+)/CD31(-), SCA1(+)/CD31(+) and SCA1(-)/CD31(-) CSP subpopulations....

Myasthenia gravis is an autoimmune disease of the neuromuscular junction (NMJ) caused by antibodies that attack components of the postsynaptic membrane, impair neuromuscular transmission, and lead to weakness and fatigue of skeletal muscle. This can be generalised or localised to certain muscle groups, and involvement of the bulbar and respiratory...

We investigated the influence of postsynaptic tyrosine kinase signaling in a mouse model of muscle-specific kinase (MuSK) myasthenia gravis (MG). Mice administered repeated daily injections of IgG from MuSK MG patients developed impaired neuromuscular transmission due to progressive loss of acetylcholine receptor (AChR) from the postsynaptic membra...

Figure S1. Percentage of MuSK‐EGFP‐positive endplates increased with the dose of rAAV injected. The tibialis anterior muscle was injected with the indicated number of viral genomes (vg) of rAAV that encoded MuSK‐EGFP. The percentage of AChR‐stained endplates that were positive for MuSK‐EGFP fluorescence was counted from sampled images. Figure S2....

Myasthenia gravis is a muscle weakness disease characterized by autoantibodies that target components of the neuromuscular junction, impairing synaptic transmission. The most common form of myasthenia gravis involves antibodies that bind the nicotinic acetylcholine receptors in the postsynaptic membrane. Many of the remaining cases are due to antib...

The neuromuscular junction (NMJ) is the large, cholinergic relay synapse through which mammalian motor neurons control voluntary muscle contraction. Structural changes at the NMJ can result in neurotransmission failure, resulting in weakness, atrophy and even death of the muscle fiber. Many studies have investigated how genetic modifications or dis...

Muscle-specific tyrosine kinase (MuSK) autoantibodies are the hallmark of a form of myasthenia gravis (MG) that can challenge the neurologist and the experimentalist. The clinical disease can be difficult to treat effectively. MuSK autoantibodies affect the neuromuscular junction in several ways. When added to muscle cells in culture, MuSK antibodi...

Purpose of review: Antibodies to muscle-specific tyrosine kinase (MuSK) characterize up to 5% of myasthenia gravis patients. This review focuses on the differences to clinical antiacetylcholine receptor-myasthenia gravis, and on the physiology and animal studies that elucidate the role of MuSK and help explain the clinical disease. Recent finding...

Key points Myasthenic anti‐muscle‐specific‐kinase (MuSK) IgG was injected into mice to study its effect upon the MuSK signalling pathway and the homeostasis of postsynaptic acetylcholine receptor packing at the neuromuscular junction. Densities of MuSK, activated Src kinase, phosphorylated ACh receptors and rapsyn were all reduced at motor endplate...

The β2-adrenergic receptor agonist, albuterol, has been reported beneficial in treating several forms of congenital myasthenia. Here, for the first time, we examined the potential benefit of albuterol in a mouse model of anti-Muscle Specific Kinase (MuSK) myasthenia gravis. Mice received 15 daily injections of IgG from anti-MuSK positive patients,...

Loss of connections between motor neurons and skeletal muscle fibers contribute to motor impairment in old age, but the sequence of age-associated changes that precede loss of the neuromuscular synapse remains uncertain. Here we determine changes in the size of neuromuscular synapses within the tibialis anterior muscle across the life span of C57BL...

Key points A mouse model of anti‐muscle‐specific kinase (MuSK) myasthenia gravis was used to study the effect of pyridostigmine (a cholinesterase inhibitor drug commonly used in myasthenia) on the disease process at the neuromuscular junction. In mice receiving injections of anti‐MuSK‐positive patient IgG, pyridostigmine treatment for 7–9 days did...

Ischemic heart disease is a major cause of morbidity and mortality worldwide. Stem cell-based therapy, which aims to restore cardiac structure and function by regeneration of functional myocardium, has recently been proposed as a novel alternative treatment modality. Resident cardiac stem cells (CSCs) in adult hearts are a key cell type under inves...

In myasthenia gravis muscle weakness is caused by autoantibodies against components of the neuromuscular junction. Patient autoantibodies against muscle specific kinase (MuSK) deplete MuSK from the postsynaptic membrane and reproduce signs of myasthenia gravis when injected into mice. Here we have examined the time-course of structural and function...

At neuromuscular synapses, neural agrin (n-agrin) stabilizes embryonic postsynaptic acetylcholine receptor (AChR) clusters by signalling through the muscle-specific kinase (MuSK) complex. Live imaging of cultured myotubes showed that the formation and disassembly of primitive AChR clusters is a dynamic and reversible process favoured by n-agrin, an...

Caveolae are invaginations of the plasma membrane that are formed by caveolins. Caveolar membranes are also enriched in cholesterol, glycosphingolipids and signaling enzymes such as Src kinase. Here we investigate the effect of cell stretch upon caveolar dynamics and signaling. Transfection of C2 myoblasts with caveolin-3-YFP led to the formation o...

During the development of the synapse, agrin and neuregulin act to build acetylcholine receptor (AChR) clusters in the postsynaptic membrane. Agrin drives AChR aggregation, while neuregulin induces AChR subunit transcription. Whilst the signalling pathways of agrin and neuregulin are distinct, we describe that neuregulin may act post-translationall...

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At the neuromuscular synapse, agrin and neuregulin are released from the motor nerve terminal to trigger the formation and clustering of post-synaptic acetylcholine receptors (AChRs). Agrin acts via muscle specific kinase (MuSK) to drive the clustering of pre-existing and newly inserted AChRs in the postsynaptic membrane, a pivotal step in synapse...

Muscle Specific Kinase (MuSK) is a transmembrane tyrosine kinase vital for forming and maintaining the mammalian neuromuscular junction (NMJ: the synapse between motor nerve and skeletal muscle). MuSK expression switches on during skeletal muscle differentiation. MuSK then becomes restricted to the postsynaptic membrane of the NMJ, where it functio...

The postsynaptic muscle-specific kinase (MuSK) coordinates formation of the neuromuscular junction (NMJ) during embryonic development. Here we have studied the effects of MuSK autoantibodies upon the NMJ in adult mice. Daily injections of IgG from four MuSK autoantibody-positive myasthenia gravis patients (MuSK IgG; 45 mg day(1)i.p. for 14 days) ca...

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Published in Journal of the Peripheral Nervous System 14 Suppl. 2

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Transient receptor potential canonical 1 (TRPC1), a widely expressed calcium (Ca(2+))-permeable channel, is potentially involved in the pathogenesis of Duchenne muscular dystrophy (DMD). Ca(2+) influx through stretch-activated channels, possibly formed by TRPC1, induces muscle-cell damage in the mdx mouse, an animal model of DMD. In this study, we...

Fluorescence resonance energy transfer (FRET) experiments at neuromuscular junctions in the mouse tibialis anterior muscle show that postsynaptic acetylcholine receptors (AChRs) become more tightly packed during the first month of postnatal development. Here, we report that the packing of AChRs into postsynaptic aggregates was reduced in 4-week pos...

A subset of myasthenia gravis patients that are seronegative for anti-acetylcholine receptor (anti-AChR) antibodies are instead seropositive for antibodies against the muscle-specific kinase (anti-MuSK-positive). Here, we test whether transfer of IgG from anti-MuSK-positive patients to mice confers impairment of the neuromuscular junction and muscl...

ASMR Oral Presentation

Neuromuscular synaptic transmission depends upon tight packing of acetylcholine receptors (AChRs) into postsynaptic AChR aggregates, but not all postsynaptic AChRs are aggregated. Here we describe a new confocal Fluorescence Resonance Energy Transfer (FRET) assay for semi-quantitative comparison of the degree to which AChRs are aggregated at synaps...

We have eyewitness testimony from scores of the participants in the four voyages of Christopher Columbus as they gave depositions in the famous series of lawsuits between the Spanish crown and the Columbus family. Through these accounts, we can see a slowly developing picture of the lands and shores the witnesses had seen. We can observe the ceremo...

Neural agrin is a heparan sulphate proteoglycan first defined by its ability to induce the clustering of acetylcholine receptors (AChRs) on cultured muscle cells. Neural agrin activates the transmembrane Muscle Specific Kinase (MuSK) on the postsynaptic muscle cell to stabilise the developing neuromuscular synapse. Three biological mechanisms for a...

We describe the design and implementation of a double-well optical lattice suitable for isolating and manipulating individual pairs of atoms. This lattice will be used to test controlled atom motion and controlled two-qubit gates.

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Using four coherent laser beams we make two different 2-D optical lattices with polarization-dependent periodicity. The lattices are coincident, and by shifting one with respect to the other, we create an array of double-well potentials.

The metabolic turnover of nicotinic ACh receptors (AChR) at the neuromuscular synapse is regulated over a tenfold range by innervation status, muscle electrical activity and neural agrin, but the downstream effector of such changes has not been defined. The AChR-associated protein rapsyn is essential for forming AChR clusters during development. He...

Agrin and neuregulin (HRG-beta1) play complementary roles in synapse formation. While HRG-beta1 induces transcriptional up-regulation of postsynaptic proteins, here we present evidence that it can potentiate agrin-induced acetylcholine receptor (AChR) clustering in C2 myotubes. Agrin induced maximal AChR clustering in 4 h. HRG-beta1 treatment for 4...

We experimentally study the superfluid-to-Mott-insulator (SF-MI) quantum phase transition in an interacting 1-d atomic gas. We realize a 1-d Bose gas by partitioning a magnetically-trapped ^87Rb Bose-Einstein condensate into an array of independent ``tubes'' formed by a deep 2-d optical lattice potential. The radial degree of freedom is frozen-out...

Contraction of the smooth muscle in the mouse vas deferens is elicited by sympathetic nerves releasing at least two neurotransmitters, adenosine triphosphate (ATP) and noradrenaline (NA). Several studies have indicated the presence of regional variation in the purinergic and noradrenergic contributions to sympathetic nerve-evoked contractions in ro...

Coyle et al. (2002), in this issue of Neuron, reveal the crystal structure for the GABA(A) receptor binding protein, GABARAP. They show GABARAP can switch from a monomer to an extended linear polymer form that may function to assemble microtubules during the intracellular trafficking or postsynaptic clustering of GABA(A) receptors.

The multiple causes and multiple consequences of mammalian heart failure make it an attractive proposition for analysis using gene array technology, especially where the failure is idiopathic in nature. However, gene arrays also hold potential artefacts, particularly when gene expression levels are low, and where changes in expression levels are mo...

P2X1-type purinoceptors have been shown to mediate fast transmission between sympathetic varicosities and smooth muscle cells in the mouse vas deferens but the spatial organization of these receptors on the smooth muscle cells remains inconclusive. Voltage clamp techniques were used to estimate the amplitudes of spontaneous excitatory junction curr...

P2X(1) receptors are ATP-gated cation channels that mediate the fast, purinergic component of sympathetic nerve-smooth muscle neurotransmission in the mouse vas deferens and may serve comparable functions in the urinary bladder and the arteries. The gene for mouse P2X(1) (P2rx1) was cloned and its genomic structure defined by sequencing. The gene s...

ATP released by sympathetic varicosities of the mouse vas deferens binds to P2X receptors which activate fast, ligand-gated channels, resulting in depolarisation of smooth muscle cells. We examined the development of fast neuromuscular transmission at surface longitudinal smooth muscle fibres of the mouse vas deferens. Sympathetic varicosities were...

Rapsyn is a protein that interacts with the cytoplasmic face of the nicotinic acetylcholine receptors (AChR) to cluster them within postsynaptic membrane of muscle. Here we show that intracellular AChRs are also affected by rapsyn. When rapsyn was co-transfected with AChR into QT-6 fibroblasts, (125)I-alpha-bungarotoxin binding indicated a reductio...

Rapsyn is a protein on the cytoplasmic face of the postsynaptic membrane of skeletal muscle that is essential for clustering acetylcholine receptors (AChR). Here we show that transfection of rapsyn cDNA can restore AChR clustering function to muscle cells cultured from rapsyn deficient (KORAP) mice. KORAP myotubes displayed no AChR aggregates befor...

The expression and subcellular localisation of dynamin and syntaxin were examined during the periods of motor neuron development and neuromuscular synaptogenesis in the mouse embryo. Both dynamin and syntaxin could be detected by immunoblotting in the spinal cord at embryonic day 10 (E10; 2 days before axon outgrowth) and at all subsequent ages exa...

Rapsyn is a 43-kDa cytoplasmic protein that clusters nicotinic acetylcholine receptors (AChR) in the postsynaptic membrane. Here we examine the effect of rapsynmediated AChR clustering on the metabolic stability of the AChR. When transfected into QT-6 fibroblasts, cell surface AChRs (alpha, beta, epsilon, and delta subunit combination) pulse labele...

Rapsyn, a 43-kDa protein on the cytoplasmic face of the postsynaptic membrane, is essential for clustering acetylcholine receptors (AChR) at the neuromuscular junction. When transfected into nonmuscle cells (QT-6), rapsyn forms discrete membrane domains and can cluster AChR into these same domains. Here we examined whether rapsyn can cluster other...

1. The post-synaptic membranes of neurons and muscle cells are characterized by clusters of transmitter receptors, the number and type of which help to determine synaptic efficacy. Here I briefly review what is known of the mechanism of clustering of nicotinic acetylcholine receptors (AChR) at neuromuscular synapses. 2. The extracellular protein ag...

Recombinant acetylcholine receptors (AChRs) expressed on the surface of cultured fibroblasts become organized into discrete membrane domains when the 43-kD postsynaptic protein (43k) is co-expressed in the same cells (Froehner, S.C., C. W. Luetje, P. B. Scotland, and J. Patrick, 1990. Neuron. 5:403-410; Phillips, W. D., M. C. Kopta, P. Blount, P. D...

The 43K protein is a cytoplasmic peripheral membrane protein concentrated subsynaptically in skeletal muscle. Recombinant 43K has been shown to cause clustering of acetylcholine receptors (AChRs) in cultured cells. However, the role of 43K in vivo is disputed, because in some cases it appears only after AChRs have clustered. We therefore examined t...

The developing neuromuscular junction has provided an important paradigm for studying synapse formation. An outstanding feature of neuromuscular differentiation is the aggregation of acetylcholine receptors (AChRs) at high density in the postsynaptic membrane. While AChR aggregation is generally believed to be induced by the nerve, the mechanisms u...

The postsynaptic membrane of the neuromuscular junction contains a myristoylated 43-kD protein (43k) that is closely associated with the cytoplasmic face of the nicotinic acetylcholine receptor (AChR)-rich plasma membrane. Previously, we described fibroblast cell lines expressing recombinant AChRs. Transfection of these cell lines with 43k was nece...

Neurotransmitter receptors are generally clustered in the postsynaptic membrane. The mechanism of clustering was analyzed with fibroblast cell lines that were stably transfected with the four subunits for fetal (alpha, beta, gamma, delta) or adult (alpha, beta, epsilon, delta) type mouse muscle nicotinic acetylcholine receptors (AChRs). Immunofluor...

The spatial distribution of intracellular acetylcholine receptors along the length of fibres from the avian posterior latissimus dorsi muscle has been investigated during embryonic development, when distributed synaptic sites are eliminated from the muscle fibres. Cell surface AChR were irreversibly blocked with unlabelled α-bungarotoxin (α-BGT). M...

Changes in the distribution of large acetylcholine receptor clusters (AChR-Cs) on developing fast-twitch fibres of the chicken posterior latissimus dorsi (PLD) muscle have been studied during the period of loss of polyneuronal innervation using fluorescein-conjugated alpha-bungarotoxin. Embryonic muscles were ultrasonically dissociated into single...

The development of the focal localization of large acetylcholine receptor clusters (AChR-Cs) on avian fast muscle fibres has been investigated in the triceps brachii pars humeralis (TH) muscle of the chick embryo. The mature TH muscle consists of both fast fibres, which usually receive a focal innervation at single synaptic sites, and slow fibres w...

Many avian muscles contain a characteristic topographical distribution of fibre types. In order to study the role of nerves in the establishment and distribution of these fibre types, monoclonal antibodies (McAb) to the heavy chain subunit of myosin (MHC) were produced. The anti-fast McAb (2B12) bound to adult fast MHC and cross-reacted with the em...

The size and distribution of acetylcholine receptor clusters (AChR-C) on normal and aneural developing muscle fibres of the chick wing were studied by labelling AChR with fluorescent conjugates of alpha-bungarotoxin (alpha-BGT). AChR-C of a size typical of initial synaptic contacts (5 micron long) were present at 7 days incubation, shortly after th...

The embryonic precursors of the avian slow (type I and III) and fast (type II) fibers can be distinguished from each other early in muscle formation (stage 28, V. Hamburger and H. L. Hamilton, J. Morphol, 88, 49-92, 1951) on the basis of the differential sensitivity of their myosin ATPases. To test the neural dependence of fiber type differentiatio...