William K. Ovalle

Functional regeneration in the hindlimb skeletal muscle of the mdx mouse.

SummaryThe pattern of spontaneous skeletal muscle degeneration and clinical recovery in hindlimb muscles of the mdx mutant mouse was examined for functional and metabolic confirmation of apparent structural regeneration. The contractile properties, histochemical staining and myosin light chain and parvalbumin contents of extensor digitorum longus (EDL) and soleus (Sol) muscles of mdx and age-matched control mice were studied at 3–4 and 32 weeks. Histochemical staining (myofibrillar ATPase and NADH-tetrazolium reductase) revealed no significant change in slow-twitch-oxidative (SO) or fast-twitch-oxidative-glycolytic (FOG) fibre type proportions in mdx Sol apart from the normal age-related increase in SO fibres. At 32 weeks mdx EDL, however, showed significantly smaller fast-twitch-glycolytic (FG) and larger FOG proportions than those in control EDL. These fibre type distributions were confirmed by differential staining with antibodies to myosin slow-twitch and fast-twitch heavy chain isozymes. Frequency distribution of cross-sectional area for each fibre type showed a wider than normal range of areas especially in FOG fibres of mdx Sol, and FG fibres of mdx EDL, supporting previous observations using autoradiography of myofibre regeneration. Isometric twitch and tetanic tensions in Sol were significantly less than in controls at 4 weeks, but by 32 weeks, values were not different from age-matched controls. In mdx EDL at 3 weeks, twitch and tetanus tensions were significantly less, and time-to-peak twitch tensions were significantly faster than in control EDL. By 32 weeks, mdx EDL twitch and tetanus tensions expressed relative to muscle weight continued to be significantly lower than in age-matched controls, despite normal absolute tensions. The maximum velocity of shortening in 32-week mdx EDL was significantly lower than in control EDL. Myosin light chain distribution in mdx Sol exhibited significantly less light chain 2-slow (LC2s) and more light chain 1b-slow(LC1bs) at 32 weeks than age-matched control Sol. Gels of EDL from 32-week-old mdx mice showed significantly less light chain 2-fast-phosphorylated (LC2f-P) and light chain 3-fast (LC3f) and significantly more light chain 1-fast (LC1f) and light chain 2-fast (LC2f), but normal parvalbumin content compared to age-matched controls. These observations suggest that mdx hindlimb muscles are differentially affected by the disease process as it occurs in murine models of dystrophy. However, the uniqueness of mdx Sol and to a lesser extent EDL is that they also undergo an important degree of functional regeneration which is able to compensate spontaneously for degenerative influences of genetic origin. The mdx mutant may therefore be an important model for the study of regeneration by skeletal muscle, and of the nerve-muscle interactions which enable or restrict that regeneration.

Morphological study of two human facial muscles: Orbicularis oculi and corrugator supercilii

Human facial muscles are unique in that they do not cross joints and they function either to open and close the apertures of the face or to tug the skin into intricate movements producing facial expressions. Compared to other skeletal muscles of the body, little is known about the microscopic architecture and organization of facial muscles. It was hypothesized that facial muscles with different roles would possess differences in their cellular organization and morphology that would reflect their unique function. The palpebral orbicularis oculi (oo) and the corrugator supercilii (cs) were studied because they are in close topographical proximity to one another and share the same nerve supply and embryonic origin. This study compared the two muscles which were procured as biopsies from cosmetic surgery procedures. Architectural and morphological features were elucidated using a combination of conventional histological stains, immunocytochemistry and histochemistry. Quantitative measures of fiber sizes, shapes, and fiber‐type distributions were performed along with measures of capillary area per unit of contractile area (capillary index). Fiber‐type profiles and motor end‐plates were demonstrated by using antibodies to fast and slow myosins, as well as to neurofilament protein. The oo was shown to differ significantly from the cs on the basis of fiber shapes, sizes, and types. The oo muscle fibers were small, rounded, and 89% of them were of the fast‐twitch (Type II) variety. The muscle fibers in the cs were larger, polygonal, and only 49% of them were of the fast‐twitch variety. The capillary index of the cs was 2.4 times that of the oo. Clin. Anat. 12:1–11, 1999.

Quantitative morphology of mast cells in skeletal muscle of normal and genetically dystrophic mice

Mast cells are indigenous connective tissue cells that function in the process of inflammation and edema. Their numbers were studied in a quantitative morphological study of the soleus muscles from 32‐week‐old and 56‐week‐old normal and genetically dystrophic dy2J and mdx mice to determine the incidence of mast cells in muscle to increasing age and to normal and myopathic conditions.

A new paradigm for teaching Histology laboratories in Canada's first distributed medical school

To address the critical problem of inadequate physician supply in rural British Columbia, The University of British Columbia (UBC) launched an innovative, expanded and distributed medical program in 2004–2005. Medical students engage in a common curriculum at three geographically distinct sites across B.C.: in Vancouver, Prince George and Victoria. The distribution of the core Histology course required a thorough revision of our instructional methodology. We here report our progress and address the question “How does one successfully distribute Histology teaching to remote sites while maintaining the highest of educational standards?” The experience at UBC points to three specific challenges in developing a distributed Histology curriculum: (i) ensuring equitable student access to high quality histological images, (ii) designing and implementing a reliable, state‐of‐the‐art technological infrastructure that allows for real‐time teaching and interactivity across geographically separate sites and (iii) ensuring continued student access to faculty content expertise. High quality images—available through any internet connection—are provided within a new virtual slide box library of 300 light microscopic and 190 electron microscopic images. Our technological needs are met through a robust and reliable videoconference system that allows for live, simultaneous communication of audio/visual materials across the three sites. This system also ensures student access to faculty content expertise during all didactic teaching sessions. Student examination results and surveys demonstrate that the distribution of our Histology curriculum has been successful. Anat Sci Ed 1:95–101, 2008.

Changes in isometric contractile properties of fast-twitch and slow-twitch skeletal muscle of dystrophic mice during postnatal development

Abstract Our primary aim was to determine if there exists a preferential involvement of the fast-twitch or slow-twitch skeletal muscle fibers in the dy 2J dy 2J strain of murine dystrophy. The changes in the contractile properties of the slow-twitch soleus (SOL) and the fast-twitch extensor digitorum longus (EDL) muscles of normal and dystrophic mice were studied at 4, 8, 12, and 32 weeks of age. Isometric twitch and tetanus tension were decreased in the 4- and 8-week-old dystrophic EDL compared with controls, this situation being reversed in the older animals. At 12 weeks, the dystrophic EDL generated 15% more tetanic tension than normal EDL and by 32 weeks no significant difference was seen between normal and dystrophic EDL twitch or tetanus tension. By 8 weeks, dystrophic EDL exhibited a prolonged time-to-peak twitch tension (TTP) and half-relaxation time ( 1 2 RT ) of the isometric twitch which continued to 32 weeks. For the dystrophic SOL, decreased twitch and tetanus tension was observed from 4 to 32 weeks. At 8 and 12 weeks, TTP and 1 2 RT of dystrophic SOL were prolonged. However, by 32 weeks there was no longer a significant difference seen in TTP or 1 2 RT between normal and dystrophic SOL. Our results appear to indicate that a loss of the primary control which is determining the fiber composition of the individual muscles is occurring as the dystrophic process advances.

STRUCTURE, DISTRIBUTION AND INNERVATION OF MUSCLE SPINDLES IN AVIAN FAST AND SLOW SKELETAL MUSCLE

Muscle spindles in 2 synergistic avian skeletal muscles, the anterior (ALD) and posterior (PLD) latissimus dorsi, were studied by light and electron microscopy to determine whether morphological or quantitative differences existed between these sensory receptors. Differences were found in the density, distribution and location of muscle spindles in the 2 muscles. They also differed with respect to the morphology of their capsules and intracapsular components. The slow ALD possessed muscle spindles which were evenly distributed throughout the muscle, whereas in the fast PLD they were mainly concentrated around the single nerve entry point into the muscle. The muscle spindle index (number of spindles per gram wet muscle weight) in the ALD was more than double that of its fast‐twitch PLD counterpart (130.5±2.0 vs 55.4±2.0 respectively, n=6). The number of intrafusal fibres per spindle ranged from 1 to 8 in the ALD and 2 to 9 in the PLD, and their diameters varied from 5.0 to 16.0 μm and 4.5 to 18.5 μm, respectively. Large diameter intrafusal fibres were more frequently encountered in spindles of the PLD. Unique to the ALD was the presence of monofibre muscle spindles (12.7% of total spindles observed in ALD) which contained a solitary intrafusal fibre. In muscle spindles of both the ALD and PLD, sensory nerve endings terminated in a spiral fashion on the intrafusal fibres in their equatorial regions. Motor innervation was restricted to either juxtaequatorial or polar regions of the intrafusal fibres. Outer capsule components were extensive in polar and juxtaequatorial regions of ALD spindles, whereas inner capsule cells of PLD spindles were more numerous in juxtaequatorial and equatorial regions. Overall, muscle spindles of the PLD exhibited greater complexity with respect to the number of intrafusal fibres per spindle, range of intrafusal fibre diameters and development of their inner capsules. It is postulated that the differences in muscle spindle density and structure observed in this study reflect the function of the muscles in which they reside.

Twelve tips for teaching in a provincially distributed medical education program.

Background: As distributed undergraduate and postgraduate medical education becomes more common, the challenges with the teaching and learning process also increase. Aim: To collaboratively engage front line teachers in improving teaching in a distributed medical program. Method: We recently conducted a contest on teaching tips in a provincially distributed medical education program and received entries from faculty and resident teachers. Results: Tips that are helpful for teaching around clinical cases at distributed teaching sites include: ask “what if ” questions to maximize clinical teaching opportunities, try the 5-min short snapper, multitask to allow direct observation, create dedicated time for feedback, there are really no stupid questions, and work with heterogeneous group of learners. Tips that are helpful for multi-site classroom teaching include: promote teacher–learner connectivity, optimize the long distance working relationship, use the reality television show model to maximize retention and captivate learners, include less teaching content if possible, tell learners what you are teaching and make it relevant and turn on the technology tap to fill the knowledge gap. Conclusion: Overall, the above-mentioned tips offered by front line teachers can be helpful in distributed medical education.

ULTRASTRUCTURE OF THE LARVAL TENTACLE AND ITS SKELETAL MUSCLE IN XENOPUS LAEVIS

During premetamorphic development, tadpoles of Xenopus laevis possess a transitory pair of long, slender, mobile tentacles situated at the corners of the mouth. Microscopic examination of the larval tentacle typically reveals three distinct compartments: a central core of cartilage, a laterally situated skeletal muscle, and a nerve supply medially. Along the length of each tentacle, the epidermis is supplied by many unmyelinated nerve fibers, presumably sensory in nature, which terminate as naked axons in close association with the epidermal cells. The striated tentacular muscle, in the proximal region of the lateral compartment, consists of extrafusal muscle fibers of varying size which range in number from 36 to 48 per tentacle (n = 10). Using morphometric criteria, we have classified the skeletal muscle fibers of the larval tentacular muscle into three types: large (30-50 microns), intermediate (20-30 microns), and small (10-20 microns). By electron microscopy, each type displays characteristic sarcomeric banding patterns, sarcotubular and mitochondrial disposition, and motor endplate ultrastructure. Our morphological observations indicate that the tentacles of the Xenopus tadpole are complex mobile facial extensions which may play roles in mechanoreception and/or chemoreception during the waterborne stages of development. Because of its transitory nature, the Xenopus tentacle may be a useful experimental model in future studies of neuromuscular development and subsequent regression in a relatively short period of time.