Despite the importance of proper synaptogenesis in the CNS, the molecular mechanisms that regulate the formation and development of synapses remain poorly understood. Indeed, the mechanisms through which initial synaptic contacts are established and modified during synaptogenesis have not been fully determined and a precise understanding of these mechanisms may shed light on synaptic development, plasticity and many CNS developmental diseases. The development and formation of spiny synapses has been thought to occur via filopodia shortening followed by the recruitment of proper postsynaptic proteins, however the precise function of filopodia remains controversial. Thus the goal of this study was to investigate the dynamics of dendritic filopodia and determine their role in the development of synaptic contacts. We initially define and characterize short lipidated motifs that are sufficient to induce process outgrowth. Indeed, the palmitoylated protein motifs of GAP-43 and paralemmin are sufficient to induce filopodial extensions in heterologous cells and to increase the number of filopodia and dendritic branches in neurons. We showed that the morphological changes induced by these FIMs (filopodia inducing motifs) require on-going protein palmitoylation and are modulated by a specific GTPase, Cdc42, that regulates actin dynamics. We also show that their function is palmitoylation dependent and is dynamically regulated by reversible protein palmitoylation. Significantly, our work suggests a general role for those palmitoylated motifs in the development of structures important for synapse formation and maturation. We combined several approaches to monitor the formation and development of filopodia. We show that filopodia continuously explore the environment and probe for appropriate contacts with presynaptic partners. We find that shortly after establishing a contact with axons, filopodia induce the recruitment of presynaptic elements. Remarkably, we find that expression of acylated motifs or the constitutively active form of cdc-42 enhances filopodia number and motility, but reduces the recruitment of synaptophysin positive presynaptic elements and the probability of forming stable axo-dendritic contacts. We provide evidence for the rapid transformation of filopodia to spines within hours of imaging live neurons and reveal potential molecules that accelerate this process.

Multi-echo T2 relaxation measurements of the human spinal cord (SC) reveal a short T2 pool of water believed to arise from water trapped between myelin bilayers, where the proportion of this water to the total water signal is called the myelin water fraction (MWF). In the present study, MWF were measured in the healthy human cervical spine at the C4-C6 vertebral levels in vivo using a 3D modified 32 echo CPMG sequence to acquire axial slices perpendicular to the cord. Volunteers were recruited in two age ranges, under 30 years old and over 50 years old, and a subset of both groups were scanned twice to test reproducibility. Mean MWF in the dorsal and lateral column WM of the group under 30 years of age was 0.29 (0.01) (mean(SE)), which agrees with previously reported MWF values in the cervical spine. The mean absolute difference between two scans was 0.06 or 26%. A negative correlation between WM MWF and age was hinted at in these findings, however more subjects are required to improve statistical power. This study paves the way for the use of 3D myelin water imaging in the cervical spine at 3.0T for the assessment of SC WM pathology.

Lower back injuries are an illness which plague our society. Although almost everyone will experience some form of lower back pain in his or her lifetime, it is a sickness that is poorly understood, calling for new and innovative research. Much of this back pain is attributed to mechanical factors. Hence, it is important to understand the mechanics of the spine and the mechanical effects of degenerative changes that may lead to back pain. The spine is a complex three-dimensional structure and it is therefore necessary to study its mechanics with in vitro tests that replicate physiological movements as closely as possible. Traditional spinal testing machines have been unable to simulate the kinematic behaviour of intervertebral joints as they have limited degrees of freedom and cannot produce dynamic motion. The first aim of this research was to commission a robotic testing facility to overcome the limitations of traditional testing machines. This facility incorporated a six degree-of-freedom (DOF) robot arm with a six DOF force transducer. Mechanical tests performed on this facility could simulate the dynamic three-dimensional kinematics of the lumbar spine. In addition, this research aimed to assess the existence of a region of laxity during spinal joint motion defined as the Neutral Zone, and to determine the effect of specific lesions introduced into the intervertebral disc. To investigate these aims, in vitro mechanical tests on spinal specimens were performed using the robotic testing facility. To ensure these tests produced experimental results that were indicative of the mechanics of the spine in life, the intervertebral disc height had to be representative of the disc height in life. A set of experiments was performed to determine a method for ensuring this. The post-mortem disc height change due to a period of time exposed to a moist environment, freezing, defrosting and application of a constant compressive load was documented in a group of sheep spines. Specimens that were frozen immediately upon removal from the body produced the most predictable results. These specimens required no preloading to ensure the disc height during mechanical testing was similar to that in life. In accordance with this result, specimens used in the ensuing mechanical tests were frozen immediately on removal from the body and stored frozen until required for testing. Tests performed on sheep spines with the robotic facility verified the existence of a Neutral Zone. A criterion was determined that defined the Neutral Zone as the region of spinal joint rotation where the gradient of the load/deformation curve is within +/-0.5 dNm/degree from zero. This definition was used to determine the extent of the Neutral Zone in spinal motion during different movements. A Neutral Zone of approximately four degrees was found in intact spinal motion segments during flexion/extension. Only spinal musculature can stabilise the spine in this region of rotation. The removal of the zygapophysial joints increased the Neutral Zone in flexion/extension by approximately two degrees and caused the appearance of a Neutral Zone in axial rotation of approximately one degree. This suggests that during these motions, the zygapophysial joints are the main passive stabilisers. The mechanical effects of intervertebral disc lesions were examined by experimentally introducing three types of tears (rim lesions, radial tears and concentric tears) into sheep intervertebral discs and comparing the mechanical response of the injured joint to the joint's response prior to the creation of the lesions. Radial tears and concentric tears had no effect on the maximum moments resisted by the intervertebral disc or the hysteresis of the joint's response to motion. An anterior rim lesion increased the Neutral Zone by approximately 1.5 degrees and reduced the maximum moment resisted by the intervertebral disc by approximately 20% during extension in L1/L2 specimens. Rim lesions were also found to reduce the maximum moment resisted by the intervertebral disc in lateral bending and axial rotation for all levels by approximately 15% and 25% respectively. Rim lesions did not affect the hysteresis of intervertebral disc motion. In summary, this research commissioned a robotic testing facility capable of simulating the dynamic, three-dimensional kinematics of the lumbar spine and provided a unique insight into the three-dimensional mechanics of intervertebral joints. Testing was performed on sheep joints, however the outcomes provide an insight into the mechanical response of the human spine. The Neutral Zone was shown to exist but only in flexion/extension. This implies that damage to the spinal muscles may produce an unstable structure during flexion/extension within this Neutral Zone. Rim lesions reduce the ability of the intervertebral disc to resist all modes of motion. This suggests that the presence of rim lesions will produce overloading of other spinal elements and instigate progressive degenerative changes.

Thesis (PhD)--University of South Australia, 2002.

Thesis (MAppSc in Physiotherapy)--University of South Australia, 1995

Thesis (M.S.)--Missouri University of Science and Technology, 2008. / Degree granted by Missouri University of Science and Technology, formerly known as University of Missouri--Rolla. Vita. The entire thesis text is included in file. Title from title screen of thesis/dissertation PDF file (viewed March 24, 2008) Includes bibliographical references (p. 87-88).

Thesis (D.M.)--University of Adelaide, 1986. / Includes bibliographical references (leaves 156-173).

Diss. (sammanfattning) Stockholm : Karol. inst., 2003. / Härtill 7 uppsatser.

Thesis (M.S.)--Ohio University, August, 1994. / Title from PDF t.p.

Thesis (Ph.D.)--University of Tennessee Health Sciences Center, 2005