Vikram Khurana Ole Isacson

Vikram Khurana and Ole Isacson led this live discussion on 29 June 2005. Readers are invited to submit additional comments by using our Comments form at the bottom of the page.

View Transcript of Live Discussion — Posted 22 August 2006

Background Text
By Vikram Khurana, M.D.
HCNR Fellow, Program in Neuroscience
Harvard Medical School

Review of concept

A universal feature of neurodegenerative diseases is the relative vulnerability of particular neuronal subpopulations. Not only do neurons secreting particular neurotransmitters appear to exhibit increased vulnerability to specific neurodegenerative diseases but, perhaps more strikingly, even within groups of neurons secreting a single neurotransmitter some groups are vulnerable while others are resistant to neurotoxic stimuli. Importantly, differential vulnerability of neurons is a feature of both sporadic and inherited forms of neurodegeneration, and is also observed in animal models of these diseases.

The pattern of neuronal death differs amongst different diseases:

In Parkinson disease (PD), whether sporadic or inherited, dopaminergic neurons of the substantia nigra pars compacta (SNPc or A9) are vulnerable, whereas other dopaminergic populations are relatively spared, including the adjacent ventral tegmental area (VTA or A10) neurons. This pattern is conserved in toxic (6-OHDA, MPTP, rotenone, proteasome inhibition) animal models of PD. Even in Drosophila, in which PD is modeled by overexpressing wild-type or mutant α-synuclein, one dopaminergic neuron group selectively degenerates (6). In Huntington disease (HD), one of several polyglutamine-associated neurodegenerative diseases, the medium spiny GABAergic projection neurons of the striatum degenerate, whereas the aspiny interneurons are spared. Intriguingly, in polyglutamine-associated diseases, the clinical phenotype and pattern of cell death becomes less distinct with increasing length of the polyglutamine tract. In amyotrophic lateral sclerosis (ALS or Lou Gehrig's disease), motor neurons do not display universal vulnerability. Some motor nuclei (III, IV, VI, and Onuf's nucleus) remain relatively intact during terminal stages of the disease, while others (V, VII, XII, and most of the spinal nuclei) usually degenerate.

In Alzheimer disease, while neuronal and synaptic loss occurs diffusely across the brain, a stereotyped spreading pattern of degeneration (entorhinal cortex à hippocampus à temporal cortexà parietofrontalcortex à basal forebrain à visual cortex) implies differential neuronal vulnerability. Indeed, even within particular regions, some neurons are more vulnerable than others (for example, CA1 is the most affected region of the hippocampus).

Understanding the basis of relative vulnerability is important. An understanding of why some neurons die and others survive when exposed to the same toxic stimulus would greatly enhance our understanding of disease pathogenesis and thereby enable more specific therapeutic strategies. For example, one could envisage pharmacologic manipulations targeted to correcting a particular "vulnerability factor" in a susceptible neuronal population. Characterization of these cell types might also allow for more refined stem cell/regenerative approaches in which transplanted cells more closely reflect lost neuronal subtype or in which cells are engineered to be more resistant to toxicity.

Defining vulnerability factors

Broadly, vulnerability of a particular neuronal population could be due to innate differences between neurons or due to surrounding factors. Innate differences: Apart from the particular neurotransmitter released by the neuron group, immunohistochemical methods have defined other differences between vulnerable and spared neurons. For example, in PD, to take only two of several differences that have been defined, vulnerable neurons in the substantia nigra are calbindin-negative and GIRK2 (G protein-coupled inwardly rectifying potassium channel 2)-positive, whereas adjacent surviving neurons in the VTA are calbindin-positive and GIRK2-negative. Innate differences may also include physical differences between neurons. For example, increased cell size has been postulated to be a factor in the vulnerability of large motor neurons in ALS. Interestingly, studies have described differences in susceptibility of dopaminergic neurons to MPTP between mouse strains in toxic models of PD. Investigating strain differences may therefore also provide insights into differential neuronal vulnerability. Laser-capture microdissection (LCM) is a technique that has greatly advanced our ability to characterize differences between neuron populations. In this technique individual neurons can be isolated and collection of a large number of neurons from a particular subpopulation allows for gene expression profiling. The major advance here has been the ability to capture a pure population of neurons without "contamination" from surrounding glia and neurons. Recently, LCM has been used by several groups to characterize innate differences between susceptible and spared neuronal populations. For example, studies have compared SNpc and VTA dopaminergic populations in normal rodents (1-3), MSN and interneurons in a mouse model of HD, and hippocampal CA1 and CA3 neurons in humans (4).

Surrounding factors: The differential vulnerability of a neuronal population could also be attributable to differences in cell-cell interactions (afferent or efferent neuronal connections or interactions with adjacent glial cells) or to other anatomic features such as vascular supply.

The use of Cre/Lox-P in mice and UAS/GAL4 in flies, which allow for temporally and spatially specific genetic manipulations, might prove very useful to investigate surrounding factors. Indeed, a recent study utilizing Cre/Lox-P demonstrated that pathological cell-cell interactions were crucial for developing cortical pathology in an HD mouse model (5). In flies, differential neuronal vulnerability has been described in models of spinocerebellar ataxia (6) and Parkinson disease (7).

Live Chat Discussion

We envisage a broad discussion of the subject, and welcome all to present their perspectives. Most broadly, we plan to discuss how elucidating mechanisms underlying differential vulnerability may be translated into therapeutic benefits for patients. We are fortunate to have as our invited guest Professor Ole Isacson from McLean Hospital/Harvard Medical School. The Isacson lab has recently performed two studies that raise important issues in this area. First, Chung et al. (1) performed LCM with microarray analysis to determine differential gene expression in SNpc and VTA neurons and proceeded to show that several differentially expressed genes alter vulnerability of cultured primary dopaminergic neurons to MPP+. This study raises the issue of functional validation of candidate vulnerability factors, and we plan to discuss experimental tools available to address this question. Second, Seo et al. (8) have shown that proteasomal function is inhibited in both vulnerable striatal neurons, non-striatal neurons, and also in skin fibroblasts from patients with Huntington disease, despite only striatal neurons being vulnerable in the disease. This study raises the possibility that a combination of innate neuronal factors, such as proteasomal function and mitochondrial activity, together with surrounding factors, such as BDNF availability, may interact to determine a neuron's ability to cope with a toxic insult.

References

1. Chung CY, S.H., Sonntag KC, Brooks A, Lin L, Isacson O. (2005) Cell type specific gene expression of midbrain dopaminergic neurons reveals molecules involved in their vulnerability and protection. Human Molecular Genetics, Epub ahead of print. Abstract

2. Greene, J.G., Dingledine, R. and Greenamyre, J.T. (2005) Gene expression profiling of rat midbrain dopamine neurons: implications for selective vulnerability in parkinsonism. Neurobiol Dis, 18, 19-31. Abstract

3. Grimm, J., Mueller, A., Hefti, F. and Rosenthal, A. (2004) Molecular basis for catecholaminergic neuron diversity. Proc Natl Acad Sci U S A, 101, 13891-13896. Abstract

4. Zucker, B., Luthi-Carter, R., Kama, J.A., Dunah, A.W., Stern, E.A., Fox, J.H., Standaert, D.G., Young, A.B. and Augood, S.J. (2005) Transcriptional dysregulation in striatal projection- and interneurons in a mouse model of Huntington's disease: neuronal selectivity and potential neuroprotective role of HAP1. Hum Mol Genet, 14, 179-189. Abstract

5. Gu, X., Li, C., Wei, W., Lo, V., Gong, S., Li, S.H., Iwasato, T., Itohara, S., Li, X.J., Mody, I. et al. (2005) Pathological Cell-Cell Interactions Elicited by a Neuropathogenic Form of Mutant Huntingtin Contribute to Cortical Pathogenesis in HD Mice. Neuron, 46, 433-444. Abstract

6. Ghosh, S., Feany, M.B. (2004) Comparison of pathways controlling toxicity in the eye and brain in Drosophila models of human neurodegenerative diseases. Hum Mol Genet 13, 2011-2018. Abstract

7. Feany, M.B. and Bender, W.W. (2000). A Drosophila model of Parkinson's disease. Nature, 404, 394-398. Abstract

8. Seo, H., Sonntag, K.C., and Isacson, O. (2004) Generalized brain and skin proteasome inhibition in Huntington's disease. Ann Neurol 56: 319-328. Abstract. Also see online coverage .

	Comments on Live Discussion
	Comment by:  John Breitner, ARF Advisor
	Submitted 22 August 2006  |  Permalink	Posted 22 August 2006
	My thoughts are that this as a very interesting approach. It capitalizes on our understandings of the populations of neurons that are (and are not) vulnerable to degeneration in Parkinson's disease, and uses relatively recent but now well developed methods to investigate reasons for the neurons' differential vulnerability. If only we knew which cells were (and were not) selectively vulnerable in AD we might try a similar approach. The problem is, I fear we don't. Not enough attention has been paid to this problem as the field has been engrossed with the amyloid theory of AD pathogenesis or, more recently, with inflammatory damage, unfortunately paying much less attention to the "targets" of these pathogenic processes. Perhaps your discussion will help focus discussion on this important area. View all comments by John Breitner
	Comment by:  Massimo Stefani
	Submitted 22 August 2006  |  Permalink	Posted 22 August 2006
	The theme of the differential susceptibility of different cell types to toxic amyloid aggregates is of outstanding importance. We have recently carried out research where we investigated the different impairment of cell viability in a wide panel of differing cultured cell lines exposed to toxic prefibrillar amyloid aggregates of a protein unrelated to any amyloid disease (the N-terminal domain of the prokaryotic HypF). Indeed, we found a wide range of susceptibilities, some cell lines being heavily affected and undergoing apoptosis, and others behaving apparently in a normal way. We found that such a different susceptibility might be traced back to different biochemical equipment in terms of molecular machineries aimed at counteracting derangements of the intracellular ion balance (particularly, free calcium and oxidative stress). In fact, we found significant correlations among resistance to the toxic effects of the aggregates and basal levels of calcium ATPase, intracellular ATP levels, and total antioxidant capacity. However, more intriguingly, we found that the...  Read more The theme of the differential susceptibility of different cell types to toxic amyloid aggregates is of outstanding importance. We have recently carried out research where we investigated the different impairment of cell viability in a wide panel of differing cultured cell lines exposed to toxic prefibrillar amyloid aggregates of a protein unrelated to any amyloid disease (the N-terminal domain of the prokaryotic HypF). Indeed, we found a wide range of susceptibilities, some cell lines being heavily affected and undergoing apoptosis, and others behaving apparently in a normal way. We found that such a different susceptibility might be traced back to different biochemical equipment in terms of molecular machineries aimed at counteracting derangements of the intracellular ion balance (particularly, free calcium and oxidative stress). In fact, we found significant correlations among resistance to the toxic effects of the aggregates and basal levels of calcium ATPase, intracellular ATP levels, and total antioxidant capacity. However, more intriguingly, we found that the more resistant cells were those with the highest content of cholesterol in their plasma membranes and vice versa. In particular, we found that the ability of the prefibrillar aggregates to interact with the plasma membranes inversely and significantly correlated with the content in cholesterol; we found also that the cells more heavily affected by the presence of the aggregates, those displaying the lowest level of membrane cholesterol, significantly improved their resistance upon enrichment of their membrane cholesterol content. It is currently widely accepted that toxic amyloid aggregates interact with cell components, notably cell membranes, and that such an interaction is a key event in favoring their translocation into the cell, out of the cell, or in their trafficking among cell compartments. It also appears fundamental in modifying the structural and biochemical properties of cell membranes leading to heavy modification of its permeability properties and, possibly, of its efficiency in signal transduction and other functions. I think that, on this aspect, the biochemical and structural properties of cell membranes are of fundamental importance, as is suggested by the above reported results and by several other previously reported data. For example, it has been shown that phosphatidylserine (mostly found in the inner membrane leaflet in normal cells) may be a preferential site of interaction for the assemblies sharing the toxic fold rich in β structure found in prefibrillar aggregates of several peptides and proteins; this could also explain the selective toxicity of folding variants of some proteins such as α lactalbumin, the ectodomain of CD44 and aprotinin to tumoral cells (known to expose phosphatidylserine molecules on the outer membrane surface). In conclusion, I think that the different susceptibility of differing cell types to the same toxic aggregates is a key topic in understanding the cell biology of aggregate toxicity; on this aspect, a central issue is a better knowledge of the role of the membrane lipid composition in favoring or disfavoring their interaction with the toxic assemblies triggering the chain of events eventually leading to cell death. View all comments by Massimo Stefani

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