Barry E. Kosofsky

Treatment of Late Infantile Neuronal Ceroid Lipofuscinosis by CNS Administration of a Serotype 2 Adeno-Associated Virus Expressing CLN2 cDNA

Late infantile neuronal ceroid lipofuscinosis (LINCL) is an autosomal recessive, neurodegenerative lysosomal storage disease affecting the CNS and is fatal by age 8 to 12 years. A total average dose of 2.5 10(12) particle units of an adeno-associated virus (AAV) serotype 2 vector expressing the human CLN2 cDNA (AAV2 CU h-CLN2) was administered to 12 locations in the CNS of 10 children with LINCL. In addition to safety parameters, a neurological rating scale (primary variable) and three quantitative magnetic resonance imaging (MRI) parameters (secondary variables) were used to compare the rate of neurological decline for 18 months in treated subjects compared with untreated subjects. Although there were no unexpected serious adverse events that were unequivocally attributable to the AAV2 CU hCLN2 vector, there were serious adverse effects, the etiology of which could not be determined under the conditions of the experiment. One subject died 49 days postsurgery after developing status epilepticus on day 14, but with no evidence of CNS inflammation. Four of the 10 subjects developed a mild, mostly transient, humoral response to the vector. Compared with control subjects, the measured rates of decline of all MRI parameters were slower, albeit the numbers were too small for statistical significance. Importantly, assessment of the neurologic rating scale, which was the primary outcome variable, demonstrated a significantly reduced rate of decline compared with control subjects. Although the trial is not matched, randomized, or blinded and lacked a contemporaneous placebo/sham control group, assessment of the primary outcome variable suggests a slowing of progression of LINCL in the treated children. On this basis, we propose that additional studies to assess the safety and efficacy of AAV-mediated gene therapy for LINCL are warranted.

Dentate gyrus volume is reduced before onset of plaque formation in PDAPP mice: A magnetic resonance microscopy and stereologic analysis

High-resolution magnetic resonance microscopy (MRM) was used to determine regional brain volumetric changes in a mouse model of Alzheimers disease. These transgenic (Tg) mice overexpress human mutant amyloid precursor protein (APP) V717F under control of platelet-derived growth factor promoter (PDAPP mice), and cortical and hippocampal β-amyloid (Aβ) deposits accumulate in heterozygotes after 8–10 mos. We used MRM to obtain 3D volumetric data on mouse brains imaged in their skulls to define genotype- and age-related changes. Hippocampal, cerebellar, and brain volumes and corpus callosum length were quantified in 40-, 100-, 365-, and 630-day-old mice. Measurements taken at age 100 days, before Aβ deposition, revealed a 12.3% reduction of hippocampus volume in Tg mice compared with WT controls. This reduction persisted without progression to age 21 mos. A significant 18% increase in hippocampal volume occurred between 40 and 630 days in WT mice, and no corresponding significant increase occurred in Tg mice. Cavalieri volume estimates of hippocampal subfields from 100-day-old Tg mice further localized a 28% volume deficit in the dentate gyrus. In addition, corpus callosum length was reduced by ≈25% in Tg mice at all ages analyzed. In summary, reduced hippocampal volume and corpus callosum length can be detected by MRM before Aβ deposition. We conclude that overexpression of APP and amyloid may initiate pathologic changes before the appearance of plaques, suggesting novel targets for the treatment of Alzheimers disease and further reinforcing the need for early diagnosis and treatment.

Cocaine Activation Discriminates Dopaminergic Projections by Temporal Response: An fMRI Study in Rat

We applied a sensitive new functional magnetic resonance imaging technique to identify the pattern and determinants of cocaine-induced brain activation in drug-naive rats. At doses greater than 0.1 mg/kg iv, cocaine produced robust activation throughout cortex with the largest magnitude increase in frontal neocortex. Additionally, we detected selective activation within dopamine-innervated subcortical regions including dorsomedial and ventrolateral striatum, nucleus accumbens region, and dorsal thalamus. Although dose response was similar among activated regions, temporal response differentiated regions along distinct anatomical boundaries with basal ganglia and limbic cortical structures, reaching maximum activation later than frontal neocortex. Pharmacological specificity was demonstrated by blocking cocaine-induced activation with SCH-23390, a selective D1 antagonist. Our data demonstrate the utility of fMRI to identify spatiotemporal patterns of cocaine-induced brain activation and implicate D1 dopaminergic mechanisms in acute cocaine action.

Newborn screening for Krabbe disease: the New York State model.

Krabbe disease is a rare inherited neurologic disorder affecting the central and peripheral nervous systems. The disease has four phenotypes: early infantile, later onset, adolescent, and adult. The only known treatment is hematopoietic stem cell transplantation, which is, in the early infantile form of the disease, most beneficial if performed before onset of clinical symptoms. In August 2006, New York State began screening all newborns for Krabbe disease. A rapid and accurate technique for assessing galactocerebrosidase activity and performing DNA mutation analysis had been developed. Interpreting these results was limited, however, because neither enzyme activity nor genetic mutation reliably predicts phenotype. A series of initiatives were therefore developed by a multidisciplinary group of neurologists, geneticists, metabolic pediatricians, neurodevelopmental pediatricians, and transplant physicians (the Krabbe Consortium of New York State) to enhance the effectiveness of the newborn screening program. A standardized clinical evaluation protocol was designed based on the available literature, criteria for transplantation for the early infantile phenotype were formulated, a clinical database and registry was developed, and a study of developmental and functional outcomes was instituted. This multidisciplinary standardized approach to evaluating infants who have positive results on newborn screening may serve as a model for other states as they begin the process of screening for Krabbe disease and other lysosomal storage disorders.

Cocaine-induced disturbances of corticogenesis in the developing murine brain

Exposure of fetuses to cocaine can induce behavioral disturbances and structural modifications of the CNS. Cocaine was injected into pregnant mice from the eighth day of gestation until term and the brains of their offspring were examined at the light and EM levels. Cocaine severely disturbs neocortical architecture, disrupting horizontal and vertical lamination, and inducing abnormal array of the axonal-dendritic bundles. Cocaine also alters several steps of gliogenesis. The severity of the malformations is variable, but evident in all exposed animals.

Regional sensitivity and coupling of BOLD and CBV changes during stimulation of rat brain

Functional MRI of rat brain was performed at 2 Tesla following intravenous injection of cocaine in order to 1) determine if changes in CBV and changes in BOLD signal were regionally coupled in brain parenchyma, and 2) compare the sensitivities of these imaging methods across different brain structures. Percent changes in CBV and BOLD relaxation rate were spatially and temporally coupled during this graded brain activation. The use of contrast agent increased functional sensitivity in all parenchymal brain structures, with a strong but predictable dependence on the resting‐state blood volume fraction. Magn Reson Med 45:443–447, 2001.

Transplacental Cocaine Exposure: A Mouse Model Demonstrating Neuroanatomic and Behavioral Abnormalities

Between 10% and 15% of infants born in urban America today have been exposed to cocaine in utero. Clinical studies have suggested that impairment of brain growth is the single best marker of significant prenatal cocaine exposure, and postnatal developmental compromise seen in a subset of affected children as a consequence of that exposure. We have developed an animal model, in mice, of prenatal cocaine exposure that has allowed us to dissociate the direct effects of cocaine in altering fetal development from the indirect effects associated with cocaine-induced malnutrition. We find that transplacental cocaine exposure independently impairs fetal brain and body growth and results in behavioral deficits and permanent alterations in neocortical cytoarchitecture in exposed offspring. (J Child Neurol 1994;9:234-241).

Volumetric cerebral characteristics of children exposed to opiates and other substances in utero

Morphometric cerebral characteristics were studied in children with prenatal poly-substance exposure (n=14) compared to controls (n=14) without such exposure. Ten of the substance-exposed children were born to mothers who used opiates (heroin) throughout the pregnancy. Groups were compared across 16 brain measures: cortical gray matter, cerebral white matter, hippocampus, amygdala, thalamus, accumbens area, caudate, putamen, pallidum, brainstem, cerebellar cortex, cerebellar white matter, lateral ventricles, inferior lateral ventricles, and the 3rd and 4th ventricles. In addition, continuous measurement of thickness across the entire cortical mantle was performed. Volumetric characteristics were correlated with ability and questionnaire assessments 2 years prior to scan. Compared to controls, the substance-exposed children had smaller intracranial and brain volumes, including smaller cerebral cortex, amygdala, accumbens area, putamen, pallidum, brainstem, cerebellar cortex, cerebellar white matter, and inferior lateral ventricles, and thinner cortex of the right anterior cingulate and lateral orbitofrontal cortex. Pallidum and putamen appeared especially reduced in the subgroup exposed to opiates. Only volumes of the right anterior cingulate, the right lateral orbitofrontal cortex and the accumbens area, showed some association with ability and questionnaire measures. The sample studied is rare and hence small, so conclusions cannot be drawn with certainty. Morphometric group differences were observed, but associations with previous behavioral assessment were generally weak. Some of the volumetric differences, particularly thinner cortex in part of the right lateral orbitofrontal cortex, may be moderately involved in cognitive and behavioral difficulties more frequently experienced by opiate and poly-substance-exposed children.

Neuroimaging of Children Following Prenatal Drug Exposure

Recent advances in MR-based brain imaging methods have provided unprecedented capabilities to visualize the brain. Application of these methods has allowed identification of brain structures and patterns of functional activation altered in offspring of mothers who used licit (e.g., alcohol and tobacco) and illicit (e.g., cocaine, methamphetamine, and marijuana) drugs during pregnancy. Here we review that literature, which though somewhat limited by the complexities of separating the specific effects of each drug from other confounding variables, points to sets of interconnected brain structures as being altered following prenatal exposure to drugs of abuse. In particular, dopamine-rich cortical (e.g., frontal cortex) and subcortical (e.g., basal ganglia) fetal brain structures show evidence of vulnerability to intrauterine drug exposure suggesting that during brain development drugs of abuse share a specific profile of developmental neurotoxicity. Such brain malformations may shed light on mechanisms underlying prenatal drug-induced brain injury, may serve as bio-markers of significant intrauterine drug exposure, and may additionally be predictors of subsequent neuro-developmental compromise. Wider clinical use of these research-based non-invasive methods will allow for improved diagnosis and allocation of therapeutic resources for affected infants, children, and young adults.

Distribution of locus coeruleus axons in the rat spinal cord: a combined anterograde transport and immunohistochemical study☆

The distribution of locus coeruleus axons in the rat spinal cord was studied by anterograde transport of Phaseolus vulgaris leucoagglutinin in combination with dopamine-beta-hydroxylase immunohistochemistry. Locus coeruleus axons were observed primarily in the superficial laminae of the dorsal horn. Few locus coeruleus fibers were seen in the vicinity of motoneuron pools or in the intermediolateral cell column of the thoracic spinal cord. The results of this study suggest a selective action of the coeruleo-spinal projection upon sensory inputs to the spinal cord.