Donald A. Siegel

Growth of Ectopic Dendrites on Cortical Pyramidal Neurons in Neuronal Storage Diseases Correlates with Abnormal Accumulation of GM2 Ganglioside

Abstract: Ganglioside analysis and quantitative Golgi studies of the cerebral cortex of cats with ganglioside and nonganglioside lysosomal storage diseases reveal a correlation between the amount of accumulated GM2 ganglioside and the extent of ectopic dendrite growth on cortical pyramidal neurons. This correlation was not observed with any of the other gangliosides assayed for, including GM1 ganglioside. These results suggest a specific role for GM2 ganglioside in the initiation of ectopic neurites on pyramidal cells in vivo and are consistent with the developing hypothesis that different gangliosides have specific roles in different cell types dependent upon the receptor or other effector molecules with which they may interact.

GM2 ganglioside and pyramidal neuron dendritogenesis

GM2 ganglioside, although scarce in normal adult brain, is the predominant ganglioside accumulating in several types of lysosomal disorders, most notably Tay-Sachs disease. Pyramidal neurons of cerebral cortex in Tay-Sachs, as well as many other types of neuronal storage disorders, are known to exhibit a phenomenon believed unique to storage disorders: growth of ectopic dendrites. Recent studies have shown that a common metabolic abnormality shared by storage diseases with ectopic dendrite growth is the abnormal accumulation of GM2 ganglioside. The correlation between increased levels of GM2 and the presence of ectopic dendrites has been found in both ganglioside and nonganglioside storage disorders, the latter including sphingomyelin-cholesterol lipidosis, mucopolysaccharidosis, and α-mannosidosis. Quantitative HPTLC analysis has shown that increases in GM2 occur in proportion to the incidence of ectopic dendrite growth, whereas, other gangliosides, including GM1, lack similar increases. Immunocytochemical studies of all nonganglioside storage diseases which exhibit ectopic dendritogenesis have revealed heightened GM2 ganglioside-immunoreactivity in the cortical pyramidal cell population, whereas neurons in normal adult brain exhibit little or no staining for this ganglioside. Further, studies examining disease development have consistently shown that accumulation of GM2 gangliosideprecedes growth of ectopic dendrites, indicating that it is not simply occurring secondary to new membrane production. These findings have prompted an examination for a similar relationship between GM2 ganglioside and dendritogenesis in cortical neurons of normal developing brain. Results show that GM2 ganglioside-immunoreactivity is consistently elevated in immature neurons during the period when they are undergoing active dendritic initiation, but this staining diminishes dramatically as the dendritic tress of these cells mature. Collectively, these studies on diseased and normal brain offer compelling evidence that GM2 ganglioside plays a pivotal role in the regulation of dendritogenesis in cortical pyramidal neurons.

Glycosylceramide synthesis in the developing spinal cord and kidney of the twitcher mouse, an enzymatically authentic model of human krabbe disease.

Abstract: UDP‐galactose:ceramide galactosyltransferase activity was assayed in the spinal cord and kidney of the recently discovered neurological mutant, the twitcher mouse, which is an enzymatically authentic model of human globoid cell leukodystrophy (Krabbe disease). The activity in the spinal cord was essentially normal during the early myelination period up to 15 days. There was a slight reduction at 20 days. At 25 and 33 days, the galactosyltransferase activity was drastically reduced compared to controls. In contrast, the galactosyltransferase activity in the kidney of twitcher mice remained normal throughout the developmental stages examined. Activity of the control enzyme UDP‐glucose:ceramide glucosyltransferase was always normal in both the spinal cord and kidney. Thus, reduction of galactosylceramide synthesis occurs in the CNS secondarily to the pathological alteration of the oligodendroglia. No such reduction occurs in the kidney, at least for the last step of galactosylceramide synthesis. Reduced synthesis as the result of metabolic regulation in the presence of the catabolic block is therefore unlikely to be the cause of the lack of abnormal accumulation of galactosylceramide in the kidney of patients with globoid cell leukodystrophy.

Ectopic axon hillock-associated neurite growth is maintained in metabolically reversed swainsonine-induced neuronal storage disease *

An experimentally induced and reversible model of a neuronal storage disease, swainsonine-induced feline alpha-mannosidosis, has been used to study the modifiability of ectopic, axon hillock-associated neurites and their new synaptic contacts. Earlier studies have fully documented that a variety of neuronal storage disorders are characterized by such changes in neuronal geometry and connectivity. Swainsonine administration was ended after 6 months of continuous treatment which had resulted in characteristic signs of alpha-mannosidosis. Studies of this animal 6 months after reversal showed that even though neuronal vacuolation and other CNS changes essentially normalized, ectopic neurites and their synaptic connections were still present and appeared similar to those of another animal which had been treated with swainsonine for the entire 12-month period. These results suggest that once initiated during the disease process, ectopic axon hillock-associated dendrites become an integral part of the soma-dendritic domain of affected neurons and may not be reversible. These findings may have relevance for current attempts to devise therapies involving enzyme replacement for individuals with inherited neuronal storage disease.

Ectopic dendrite initiation: CNS pathogenesis as a model of CNS development.

The neuronal storage diseases are a rare group of disorders with profound clinical consequences including severe mental retardation and death in early childhood. A subset of these disorders, those with elevated levels of GM2 ganglioside, are further characterized by the reinitiation of primary dendrites on mature cortical neurons. These ectopic dendrites are unusual as primary dendrite initiation is normally confined to a narrow developmental window. Thus, ectopic dendritogenesis appears to be a recapitulation of the normal developmental program temporally displaced. Consequently, understanding ectopic dendritogenesis should offer insights into both the pathogenesis of the neuronal storage diseases as well as mechanisms of normal CNS development.

Galactosylceramide synthesis in the peripheral nerve of normal and Quaking mice

Abstract The activity of UDP-galactose-ceramide galactosyltransferase, which catalyzes the last synthetic step of the myelin-characteristic lipid, galactosylceramide, was determined in the sciatic nerve of normal and Quaking mice between the ages of 5 and 40 days, and the results compared with those from brain microsomes. The newly developed assay procedure that utilized acceptor-incorporated liposomes, permitted reliable assays on very small amounts of tissues. Homogenates of the normal mouse sciatic nerve showed surprisingly high activities of the enzyme, with the specific activities similar to or higher than those of the brain microsomal fractions. The normal developmental changes were similar to those in the brain, except that the entire curve was shifted to younger ages with the peak activity occurring at 15 days. The activity in the Quaking nerve was moderately but significantly decreased already at 5 days. The difference became greater at older ages, reaching 20% of normal. This observation was similar to that in Quaking brain, except that the abnormally low activity was evident at 5 days when no significant difference could be found between normal and Quaking brains. Positive diagnosis of affected mice was possible even at 5 days by light and electron microscopic examinations. The developmental changes in the normal sciatic nerve are consistent with the known earlier myelination in the peripheral nervous system, and the finding on the Quaking nerve indicated that biochemical abnormalities accompany the pathological changes also in the peripheral nerves, as in the brain.

Batten Disease: A Typical Neuronal Storage Disease or a Genetic Neurodegenerative Disorder Characterized by Excitotoxicity?

Batten disease (neuronal ceroid lipofuscinosis) is an inherited neurological disorder of humans and a variety of animal species including dogs, mice, and sheep. Affected individuals appear normal at birth but later exhibit progressive neurological deterioration and death. The spectrum of clinical disease includes retarded mental development and/or dementia, blindness, motor system dysfunction, and seizures, and in late disease the latter can be intractable. The age at which clinical symptoms appear varies and infantile, late infantile, juvenile and adult-onset disease subtypes are recognized. Disease course in individuals with early-onset disease generally is rapid, whereas late-onset disorders exhibit a more protracted course. On postmortem exam, atrophy of cerebral cortex and ballooning of surviving neurons are characteristic features. The latter finding has led to classification of Batten disease as a neuronal storage disorder along with Tay-Sachs, Hurler, and related lysosomal diseases. Although the primary metabolic defect(s) in Batten disease remain unknown, recent research has established that, with the exception of infantile disease variants, a substantial portion of the intracellular storage material is a single protein, subunit c of mitochondrial ATP synthase.1 Current findings suggest that this subunit, which is encoded by nuclear DNA, is synthesized correctly and undergoes normal trafficking to mitochondria; however, its subsequent removal from mitochondria and degradation appear to be delayed.2 Why this particular mitochondrial component accumulates in cells, and whether its accumulation signals the primary metabolic defect in Batten disease, are unknown.

Erratum to “Ectopic dendrite initiation: CNS pathogenesis as a model of CNS development” [Int. J. Dev. Neurosci. 20 (2002) 373–389]

Fig. 5. Human Tristanin clone: (A) protein sequence: shaded area near the amino terminus represents part of the PR domain, bolded sequences are the 10 Zn-finger domains, lightly shaded and italicized area near the carboxy terminus represents the sequences used for preparing antibodies, and under lined sequence represents the area isolated in the original subtraction; (B) schematic representation of Tris; (C) comparison of Tris with other positive r gulatory domain member (PRDM) proteins, clearly demonstrating that Tris is a member of this family of transcription factors. Top line represents consensus sequence; when evenly divided, Tris sequences was chosen.