Sarah A. Dunlop

USE OF CORTICOSTEROIDS BY AUSTRALIAN OBSTETRICIANS: A SURVEY OF CLINICAL PRACTICE

Summary: All Fellows, Members and trainees of the Royal Australian College of Obstetricians and Gynaecologists resident in Australia (n = 1,281) received a questionnaire relating to their practice of prescribing antenatal corticosteroids. 833 (65%) responded. The key findings were that 97% of Australian obstetricians prescribe antenatal corticosteroids in the classical setting of uncomplicated early preterm labour and 85% prescribe repeated courses in those cases in which the risk of preterm birth persists or recurs; 50% of obstetricians prescribe this agent weekly in cases with persisting risk of preterm birth. Some of the prescribing practices were found to be related to the number of years since obtaining specialist qualification. In view of the widespread clinical use of repeated doses of corticosteroids revealed in this present survey, it is clear that further research is warranted to determine the possible benefits and hazards of repeated exposures of the developing fetus to this therapy.

Repeated prenatal corticosteroid administration delays myelination of the corpus callosum in fetal sheep

Glucocorticoids regulate oligodendrocyte maturation and the myelin biosynthetic pathways. Synthetic glucocorticoids, the corticosteroids have been successfully used in clinical practice as a single course to enhance lung maturation and reduce mortality and morbidity in preterm infants with no long‐term neurologic or cognitive side effects. However, a trend has arisen to use repeated courses despite an absence of safety data from clinical trials. We examined the effects of clinically appropriate, maternally administrated, repeated courses of corticosteroids on myelination of the corpus callosum using sheep as a large animal model. The corpus callosum is a major white matter tract that undergoes protracted myelination, underpins higher order cognitive processing and developmental damage to which is associated with, for example, cerebral palsy, mental retardation and attention deficit hyperactivity disorder. Pregnant ewes were given saline or betamethasone (0.5 mg/kg) at 104, 111, 118 and 124 days gestation, stages equivalent to the third trimester in humans. Lambs were delivered at 145 days (term), perfused and the corpus callosum examined light and electron microscopically. Total axon numbers were unaffected (P>0.05). However, myelination was significantly delayed. Myelinated axons were 5.7% in the experimental group and 9.2% in controls (P<0.05); conversely, unmyelinated axons were 88.3 and 83.7% (P<0.05). Myelinated axon diameter and myelin sheath thickness were also reduced (0.68 vs. 0.94 and 0.11 vs. 0.14 μm, P<0.05). Our data suggest that repeated prenatal corticosteroid administration delays myelination of the corpus callosum and that further safety data are needed to evaluate clinical practice.

Redefining the role of metallothionein within the injured brain: extracellular metallothioneins play an important role in the astrocyte-neuron response to injury

A number of intracellular proteins that are protective after brain injury are classically thought to exert their effect within the expressing cell. The astrocytic metallothioneins (MT) are one example and are thought to act via intracellular free radical scavenging and heavy metal regulation, and in particular zinc. Indeed, we have previously established that astrocytic MTs are required for successful brain healing. Here we provide evidence for a fundamentally different mode of action relying upon intercellular transfer from astrocytes to neurons, which in turn leads to uptake-dependent axonal regeneration. First, we show that MT can be detected within the extracellular fluid of the injured brain, and that cultured astrocytes are capable of actively secreting MT in a regulatable manner. Second, we identify a receptor, megalin, that mediates MT transport into neurons. Third, we directly demonstrate for the first time the transfer of MT from astrocytes to neurons over a specific time course in vitro. Finally, we show that MT is rapidly internalized via the cell bodies of retinal ganglion cells in vivo and is a powerful promoter of axonal regeneration through the inhibitory environment of the completely severed mature optic nerve. Our work suggests that the protective functions of MT in the central nervous system should be widened from a purely astrocytic focus to include extracellular and intra-neuronal roles. This unsuspected action of MT represents a novel paradigm of astrocyte-neuronal interaction after injury and may have implications for the development of MT-based therapeutic agents.

The Effects of Maternally Administered Methadone, Buprenorphine and Naltrexone on Offspring: Review of Human and Animal Data

Most women using heroin are of reproductive age with major risks for their infants. We review clinical and experimental data on fetal, neonatal and postnatal complications associated with methadone, the current “gold standard”, and compare these with more recent, but limited, data on developmental effects of buprenorphine, and naltrexone. Methadone is a µ-opioid receptor agonist and is commonly recommended for treatment of opioid dependence during pregnancy. However, it has undesired outcomes including neonatal abstinence syndrome (NAS). Animal studies also indicate detrimental effects on growth, behaviour, neuroanatomy and biochemistry, and increased perinatal mortality. Buprenorphine is a partial µ-opioid receptor agonist and a κ-opioid receptor antagonist. Clinical observations suggest that buprenorphine during pregnancy is similar to methadone on developmental measures but is potentially superior in reducing the incidence and prognosis of NAS. However, small animal studies demonstrate that low doses of buprenorphine during pregnancy and lactation lead to changes in offspring behaviour, neuroanatomy and biochemistry. Naltrexone is a non-selective opioid receptor antagonist. Although data are limited, humans treated with oral or sustained-release implantable naltrexone suggest outcomes potentially superior to those with methadone or buprenorphine. However, animal studies using oral or injectable naltrexone have shown developmental changes following exposure during pregnancy and lactation, raising concerns about its use in humans. Animal studies using chronic exposure, equivalent to clinical depot formulations, are required to evaluate safety. While each treatment is likely to have maternal advantages and disadvantages, studies are urgently required to determine which is optimal for offspring in the short and long term.

Neural development in metatherian and eutherian mammals: variation and constraint.

A model for predicting the timing of neurogenesis in mammals (Finlay and Darlington [1995] Science 268:1578–1584) is here extended to an additional five metatherian species and to a variety of other events in neural development. The timing of both the outgrowth of axonal processes and the establishment and segregation of connections proves to be as highly predictable as neurogenesis. Expressed on a logarithmic scale, late developmental events are as predictable as early ones. The fundamental order of events is the same in eutherian and metatherian animals, but there is a curvilinear relation between the event scales of the two; for metatherians, later events are slowed relative to earlier events. Furthermore, in metatherians, the timing of developmental events is more variable than in eutherians. The slowing of late developmental events in metatherians is associated with their considerably longer time to weaning compared with eutherians. J. Comp. Neurol. 411:359–368, 1999.

Erythropoietin is both neuroprotective and neuroregenerative following optic nerve transection

The cytokine hormone erythropoietin (EPO) is neuroprotective in models of brain injury and disease, and protects retinal ganglion cells (RGC) from cell death after axotomy. Here, we assessed EPOs neuroprotective properties in vivo by examining RGC survival and axon regeneration at 4 weeks following intraorbital optic nerve transection in adult rat. EPO was administered as a single intravitreal injection at the time of transection (5, 10, 25, 50 units, PBS control). Intravitreal EPO (5, 10 units) significantly increased RGC somata and axon survival between the eye and transection site. Twenty five units did not improve survival of RGC somata but did increase axon survival between the eye and transection site. In addition, a small proportion of axons penetrated the transection site and regenerated up to 1 mm into the distal nerve. In a second series, intravitreal EPO (25 units) doubled the number of RGC axons regenerating along a length of peripheral nerve grafted onto the retrobulbar optic nerve. Our in vivo evidence of both neuroregeneration and neuroprotection, taken together with the natural occurrence of EPO within the body and its ability to cross the blood-brain barrier, suggests that it offers promise as a therapeutic agent for central nerve repair.

Striking Denervation of Neuromuscular Junctions without Lumbar Motoneuron Loss in Geriatric Mouse Muscle

Reasons for the progressive age-related loss of skeletal muscle mass and function, namely sarcopenia, are complex. Few studies describe sarcopenia in mice, although this species is the mammalian model of choice for genetic intervention and development of pharmaceutical interventions for muscle degeneration. One factor, important to sarcopenia-associated neuromuscular change, is myofibre denervation. Here we describe the morphology of the neuromuscular compartment in young (3 month) compared to geriatric (29 month) old female C57Bl/6J mice. There was no significant difference in the size or number of motoneuron cell bodies at the lumbar level (L1–L5) of the spinal cord at 3 and 29 months. However, in geriatric mice, there was a striking increase (by ∼2.5 fold) in the percentage of fully denervated neuromuscular junctions (NMJs) and associated deterioration of Schwann cells in fast extensor digitorum longus (EDL), but not in slow soleus muscles. There were also distinct changes in myofibre composition of lower limb muscles (tibialis anterior (TA) and soleus) with a shift at 29 months to a faster phenotype in fast TA muscle and to a slower phenotype in slow soleus muscle. Overall, we demonstrate complex changes at the NMJ and muscle levels in geriatric mice that occur despite the maintenance of motoneuron cell bodies in the spinal cord. The challenge is to identify which components of the neuromuscular system are primarily responsible for the marked changes within the NMJ and muscle, in order to selectively target future interventions to reduce sarcopenia.

Activity-dependent plasticity : implications for recovery after spinal cord injury

Spinal cord injury causes devastating loss of function and progressive, potentially life-threatening, secondary complications. Although significant preclinical advances continue to be made in cellular and molecular therapies which promote regeneration, plasticity within remaining circuits and how it can be influenced by physical activity is evolving as a key research area. Understanding what constitutes plasticity, and how activity shapes it, has centred primarily on neurons, but evidence is emerging that activity also influences glial cells. Basic and clinical research continue to advance our knowledge of the quality and quantity of physical exercise required to improve function, while mental exercise is emerging as another avenue. Increased understanding of mechanisms driving activity-dependent plasticity will help develop rehabilitative strategies which optimise functional recovery.

Early Events of Secondary Degeneration after Partial Optic Nerve Transection: An Immunohistochemical Study

Secondary degeneration in the central nervous system involves indirect damage to neurons and glia away from the initial injury. Partial transection of the dorsal optic nerve (ON) results in precise spatial separation of the primary trauma from delayed degenerative events in ventrally placed axons and parent somata. Here we conduct an immunohistochemical survey of secondary cellular changes in and around axons and their parent retinal ganglion cell (RGC) somata during the first 3 days after a restricted, dorsal ON transection. This is before the secondary loss of RGCs and axons projecting through the uninjured, ventral portion of the ON. Within 5 min, manganese superoxide dismutase (MnSOD; a marker of oxidative stress) co-localizes within the astrocytic network across the entire profile of the ON. Secondary astrocyte hypertrophy of immunofluorescent labeling was evident from 3 h, with sustained increases in myelin basic protein immunoreactivity across the nerve by 24 h. Increases in NG-2-positive oligodendrocyte precursor cells, ED-1-positive activated microglia/macrophages, and Iba1-positive reactive resident microglia/macrophage numbers were only seen in ON vulnerable to secondary degeneration by 3 days. Changes within RGC somata exclusively vulnerable to secondary degeneration were detected at 24 h, as evidenced by increases in MnSOD immunoreactivity, followed by increases in c-jun immunoreactivity at 3 days. Treatment with the voltage-gated calcium channel blocker lomerizine did not alter any measured outcome. We conclude that oxidative stress spreading via the astrocytic network and from injured axons to parent RGC somata is an early event during secondary degeneration, and containment is likely to be required in order to prevent further damage to the nerve.

A morphometric study of the retinal ganglion cell layer and optic nerve from metamorphosis in Xenopus laevis

The number and distribution of cells in the retinal ganglion layer have been examined in Nissl-stained wholemounts during postmetamorphic life in Xenopus laevis. Total cell number at metamorphic climax, in the 3 and 4 month juveniles and in 5 year adults were 30,000-41,000, 45,000-51,000 and 71,000-82,000 respectively. Optic axon counts of 29,000, 39,000 and 58,000 at these stages suggested that most cells counted were ganglion cells and that their number doubled during this period. Cell distribution changed after metamorphosis producing nasal and temporal regions of higher density in juveniles and adults. These results are discussed in relation to retinal growth and plasticity of retinotectal connections.