 |
||||
|
||||
|
||||
|
||||
|
||||
Previous Article | Next Article
Volume 17, Number 3, Issue of February 1, 1997 pp. 1033-1045
Copyright ©1997 Society for NeuroscienceNeuropathology of Degenerative Cell Death in Caenorhabditis elegans
David H. Hall1, Guoqiang Gu2, Jaime García-Añoveros2, Lei Gong3, Martin Chalfie2, and Monica Driscoll3 1 Department of Neurosciences, Albert Einstein College of Medicine, Bronx, New York 10461, 2 Department of Biological Sciences, Columbia University, New York, New York 10027, and 3 Department of Molecular Biology and Biochemistry, Rutgers University, Piscataway, New Jersey 08855
ABSTRACT
In Caenorhabditis elegans necrosis-like neuronal death is induced by gain-of-function (gf) mutations in two genes, mec-4 and deg-1, that encode proteins similar to subunits of the vertebrate amiloride-sensitive epithelial Na+ channel. We have determined the progress of cellular pathology in dying neurons via light and electron microscopy. The first detectable abnormality is an infolding of the plasma membrane and the production of small electron-dense whorls. Later, cytoplasmic vacuoles and larger membranous whorls form, and the cell swells. More slowly, chromatin aggregates and the nucleus invaginates. Mitochondria and Golgi are not dramatically affected until the final stages of cell death when organelles, and sometimes the cells themselves, lyse. Certain cells, including some muscle cells in deg-1 animals, express the abnormal gene products and display a few membrane abnormalities but do not die. These cells either express the mutant genes at lower levels, lack other proteins needed to form inappropriately functioning channels, or are better able to compensate for the toxic effects of the channels. Overall, the ultrastructural changes in these deaths suggest that enhanced membrane cycling precedes vacuolation and cell swelling. The pathology of mec-4(gf) and deg-1(gf) cells shares features with that of genetic disorders with alterations in channel subunits, such as hypokalemic periodic paralysis in humans and the weaver mouse, and with degenerative conditions, e.g., acute excitotoxic death. The initial pathology in all of these conditions may reflect attempts by affected cells to compensate for abnormal membrane proteins or functions.
Key words: neurodegeneration; Caenorhabditis elegans; degenerin; mec-4; deg-1 neuropathology; necrosis; membrane cycling
INTRODUCTION
Wyllie et al. (1980)
suggested that cells die in two morphologically distinct patterns, apoptosis and necrosis, that are ubiquitous in Metazoans. In apoptosis, which occurs in normal development and homeostasis, the nucleus and cytoplasm condense, while cytoplasmic organelles maintain their integrity and DNA degrades into nucleosome-sized fragments. In necrosis, which commonly results from cell injury, cell membranes and organelles are disrupted, and the dying cells swell and lyse.
Programmed cell death in the nematode Caenorhabditis elegans shares morphological and molecular similarities with apoptosis in higher organisms (Robertson and Thomson, 1982
To understand how degenerin defects lead to cell death and to compare this degeneration with other types of cell death, we have followed the progression of neuronal pathology in mec-4(gf) and deg-1(gf) animals via light and electron microscopy. Degeneration seems to be initiated with enhanced endocytosis at the plasma membrane, the inferred location of the ion channels. The onset of the death depends on the dosage of the mutant gene. Although mec-4 and deg-1 are expressed in different neurons, cell death induced by mutant forms of either appears identical. Similarities between the degenerin-induced deaths and the early pathology of dominant myotonias (Engel, 1970
MATERIALS AND METHODS
Nematode strains. Nematode cultures were grown at 20°C unless otherwise noted, according to standard methods (Brenner, 1974
The otherwise wild-type ZB7 strain contains an array, bzIs3, that is integrated into the X chromosome and has ~10 copies of plasmid TU#14, which encodes the mec-4(u231) allele (Driscoll and Chalfie, 1991
We made the deg-1lacZ fusion construct TU#224 by subcloning a 4083 bp EcoRI-MscI fragment from a 6.7 kb deg-1 genomic clone (García-Añoveros et al., 1995 -galactosidase activity as described by Fire et al. (1990)
We produced strains with integrated arrays of the TU#224 deg-1lacZ fusion (uIs5, uIs6, and uIs7) as before (Mitani et al., 1993
Transgenic animals carrying extrachromosomal (uEx141 and uEx142) or integrated (uIs5, uIs6, or uIs7) arrays of the TU#224 deg-1lacZ fusion were stained for
Timing of gene expression and neurodegeneration. mec-4(u231) mutant animals were synchronized to within 3 hr of each other by allowing 100 gravid adults to lay eggs for 1 hr. Larvae were harvested for observation by differential interference contrast microscopy or were stained for
Electron microscopy. ZB7 animals were grown at 25°C and staged under differential interference contrast optics to select cells representing different time points of PVM degeneration. Although PVM is produced at the same time as AVM (Sulston and Horvitz, 1977
Light microscopy also was used to select time points during the death of PVC neurons in deg-1(u38) animals grown at 25°C. We had some difficulty identifying the earliest time points for cell degeneration, because the onset of cell swelling was less predictable than in ZB7 strain animals. After fixation and embedding as above, transverse serial sections were collected through the tail, including the preanal and lumbar ganglia. Complete serial sections were collected for the reported cells.
RESULTS
Several factors influence the onset and extent of degenerin-induced cell death
mec-4 normally is expressed only in a set of six touch receptor neurons (Mitani et al., 1993
Fig. 1. Summary of the timing of mec-4 and deg-1 expression and cell death. Times are given in hours after hatching at 20°C, although the data for the deg-1 animals were obtained at 25°C and converted to the equivalent times at 20°C. The production of the cell (PVM for the mec-4 animals and PVC for the deg-1 animals) from division of its precursor is shown by open circles. Filled circles indicate the time when the greatest number of cells expresses lacZ from mec-4 (>50%) or deg-1 (20%) fusions. X indicates the time when the most cell deaths are seen (>50% for both strains). Data for mec-4(e1611) came from Chalfie and Sulston (1981)
[View Larger Version of this Image (9K GIF file)]
The time of onset of PVM degeneration depended on the mec-4 mutation used and its copy number. The AVM and PVM cells became vacuolated and died in mec-4(e1611) animals when the animals were young adults (at ~45 hr after hatching; Chalfie and Sulston, 1981
The rapidly dying cells in the ZB7 animals did not swell as much as the dying cells in u231 animals. In EM reconstructions the diameter of ZB7 cells increased by approximately twofold, increasing the volume ~10-fold (data not shown). In contrast, the diameters of u231 cells, when viewed by differential interference contrast (DIC) optics, increased fivefold to give a volume increase of 100-fold. Presumably, at high gene dosage the toxic product more rapidly inactivated cell functions that contribute to swelling. Consistent with an accelerated time course of cell death, the degenerating cells also disappeared much sooner in ZB7 animals (<6 hr after the onset of visible swelling) than in u231 animals, where the touch cells can persist for 8 hr or longer as distended somata.
Specific gf alleles of deg-1 (the dominant mutations u38 and u529 and the recessive mutation u506) cause neuronal degeneration that is morphologically identical to that induced by mec-4(gf) alleles (Chalfie and Wolinsky, 1990 Predegeneration stage The two PVM neurons that we sectioned in the predegeneration stage differed from their neighboring lineage sisters, the SDQL cells, by having small membrane whorls at the plasma membrane. In one animal the PVM cell contained a small circle of internalized membrane, which appeared slightly more electron dense than normal plasma membrane or ER and may have contained a second circlet (Fig. 4). In the other animal the PVM cell had five small whorls (nested membranous shells) just inside the plasma membrane and two small circles of dense membrane lying exterior to the cell. Neither cell had vacuoles or cell swelling, and most organelles, including the nucleus, appeared normal. Two mitochondria in the second cell had increased density localized to their outer surface. In each case, as expected for this time in development, a single very short (0.5 and 4 µm) axonal process grew ventrally from the soma toward the ventral nerve cord.
As predicted for cell-autonomous expression of deg-1, a deg-1lacZ fusion was expressed in many neurons, the positions of which matched those of dying cells in u38 and u506 animals (Fig. 2). These cells included the presumptive ASH cells, IL1 cells, AVD cells, the AVG cell, and the PVC cells, although the latter stained weakly. Muscles also expressed the deg-1lacZ fusion. The head muscle quadrants and a muscle near the anus, presumably the anal depressor muscle, stained at all stages, whereas body wall muscles stained most strongly in newly hatched larvae. A similar staining pattern was seen with deg-1gfp fusions (data not shown). 
Fig. 2. Expression of deg-1lacZ. Animals contain the integrated array uIs6 and are homozygous for the sqt-1(sc103) mutation. A, Early L1 larva. B, Late L1 larva. C, L2 larva. D, L4 larva. Note staining (triangles) in head muscles at the front of the L4 larva. Cells are identified by their position within the animals.
[View Larger Version of this Image (82K GIF file)]
As with touch cell death in mec-4 mutants, the onset of PVC death depended on the deg-1 allele used (Fig. 1). Although we were able to see deg-1lacZ expression in PVC cells in only one of our constructs (uIs6), the peak of its expression preceded the peak onset of u38-induced deaths by ~6 hr. The PVC deaths induced by the u506 mutation, however, occurred much later at 25°C than those produced by the u38 mutation. This delay could result because the u506 product is either less toxic or less stable than the u38 product. With either possibility a greater amount of time would be needed to accumulate a lethal amount of the product. The onset of the cell death also depends on the dosage of the u38 mutation, because lower dosage results in a later onset of PVC degeneration (Chalfie and Wolinsky, 1990
Most of the deg-1lacZ-expressing cells, however, did not degenerate in u38 animals (data not shown). Several factors may contribute to cell lethality. First, some cells may make insufficient amounts of the toxic product. Some cell types seem to produce near-threshold amounts of the toxic product, because not all cells of a given type die. For example, variable numbers of IL1 cells (Chalfie and Wolinsky, 1990 Light microscopy of degenerin-induced cell death
We examined the degeneration of PVM in ~80 ZB7 animals using DIC microscopy and have divided the process into four periods. The first, or predegeneration, stage lasted 3 hr from the production of the PVM cell. In this initial stage the cells were not detectably different from wild-type cells when viewed with light microscopy (Fig. 3A). Obvious pathology is seen after this time (at ~2 hr before the L1 molt at 25°C). Although the subsequent pattern of degeneration was similar in all animals (Fig. 3), its time course was quite variable, lasting 4-20 hr from the start of the degeneration because of variations in the last of the four periods.
Fig. 3. Light microscopy of cell death in mec-4(gf) and deg-1(gf) animals. A-J, Degeneration of PVM in ZB7. A, A predegeneration normal-looking PVM cell ~2 hr after its production. This is not the same cell depicted in subsequent panels. The arrowhead points to the cell body. B, Start of degeneration (0 min, early degeneration). A small space (arrow) has begun to appear around the PVM nucleus (arrowhead). C, At 30 min. The cell shows obvious swelling; the nuclear envelope is not easily distinguished. D, At 90 min (middle degeneration). The cytoplasm clears, and the nucleus swells. E, At 120 min. The nucleus has moved to one side of the vacuolated space. F, At 160 min (10 min before the L1 lethargus, late degeneration). The cytoplasmic vacuole and nucleus (arrowhead) are smaller. G, At 190 min (during the L1 lethargus). The vacuole and nucleus are very poorly seen. H, At 7 hr. The vacuole and nucleus (arrowhead) are visible but condensed. I, At 19 hr. The nucleus (arrowhead) has condensed further. J, At 24 hr. Only debris (arrowhead) remains. K-N, Degeneration of PVC in deg-1(u38). K, At ~0.5 hr after the start of degeneration. L, At 1 hr. The nucleus has enlarged. M, At 1.5 hr. The nucleus begins to condense. N, At 2.2 hr. The nucleus continues to condense, especially at its periphery. The cell disappeared in another 2.2 hr.
[View Larger Version of this Image (121K GIF file)]
The first signs of pathology were detectable in the second, or early degeneration, stage (Fig. 3B,C). Initially, a small space, which was neither clear nor filled with particles displaying Brownian motion, appeared between the plasma membrane and the nucleus. This space enlarged rapidly so that the neuron swelled to its maximal size by the end of this period. The nucleus did not change in size during this period, but its outline was no longer clear (Fig. 3C). The early stage was extremely short, lasting ~15 min.
In the third, or middle degeneration, period, which lasted ~2 hr (Fig. 3D,E), the space between the plasma membrane and the nucleus cleared and contained particles displaying Brownian motion. The nucleus began to swell to two or three times its normal size, and it was displaced gradually to one side of the cell. The nucleus also contained particles showing Brownian motion.
The onset of the fourth, or late degeneration, stage (Fig. 3F-J) came abruptly. The vacuoles in both the cytoplasm and the nucleus shrunk very quickly, and the cell decreased in size. The nucleus gradually condensed and became a small core by the time of the L1 lethargus (the dormancy period preceding the molt). In some animals debris from the PVM cell disappeared in the first hour after the L1 molt. In others the PVM vacuole was more apparent after the molt (Fig. 3H). Sometimes a similar disappearance/reappearance occurred at the L2 lethargus. These apparent changes may be a consequence of the cells being obscured by the molting cuticle or may result from different osmotic conditions during lethargus. All cells had disappeared by the late L3 stage. (The late disappearance of the cells in many observed animals may result from the constant viewing of the cells under a coverslip, because we rarely saw debris from cells 6 hr after the L1 molt in staged animals taken directly from plates.)
The degeneration of the PVC interneurons in deg-1(u38) and deg-1(u506) mutants viewed in the light microscope progressed similarly to the death of PVM cells in ZB7 animals (Fig. 3K-N) (data not shown). Cells began to swell and vacuolate before nuclear morphology changed. Nuclei then swelled and began to accumulate refractile material. The cells were not detected 4 or 5 hr after they had begun to vacuolate. The two deg-1 mutants differed, however, in the time of onset of the deaths; the u506 deaths occurred later, although at a less precise time (see Fig. 1).
In the later stages of the deaths in mec-4(gf) and deg-1(gf) animals, dying cells appeared to condense, a characteristic found in programmed cell death. Although mutations in ced-3 and ced-4, genes that are needed for programmed cell death, do not affect the onset of mec-4(gf)- and deg-1(gf)-induced cell deaths (Ellis and Horvitz, 1986 Ultrastructure of degenerin-induced cell death
We used serial section reconstruction to examine the ultrastructure of a total of seven PVM cells and one AVM cell in the mec-4(u231) overexpression strain ZB7, two at each of the four stages defined by our light microscopic observations. The electron micrographs revealed a reproducible sequence of cellular changes that accompany degeneration.
Fig. 4. The predegeneration stage. A, Section through the somata of PVM and SDQL. In this section the cells look very similar. Note the dispersed chromatin in both nuclei (labeled with the cell names), the strands of ER in the dark cytoplasm of PVM (thin black arrows), and the characteristic ventralward process extending from PVM (open curved arrow). SDQL has a rostral process that begins to extend at this time (not shown). B, A glancing section through a more caudal portion of the same cells as in A. The thin white arrow points to a very small circular set of nested membranes that seems to be the earliest sign of mec-4(gf)-induced degeneration in the PVM cell. Organelles and dark cytoplasm in PVM and SDQL look similar.
[View Larger Version of this Image (165K GIF file)]
Early degeneration stage The two PVM neurons in the early degeneration stage showed many abnormalities (small internal membrane whorls and internal vacuoles) that may have formed from the plasma membrane, because many showed direct connections to it. Other cytoplasmic organelles, however, were still intact and unswollen. One cell had many small whorls surrounding the soma (possibly shed pieces of plasma membrane), three small cytoplasmic vacuoles, and a normal-appearing nucleus (Fig. 5A,B). The degeneration of the second cell appeared more advanced in all respects, with more and larger whorls, larger vacuoles, and early signs of nuclear degeneration (chromatin clumping along the inner surface of the nuclear envelope; Fig. 5C). A few membrane whorls also were found outside of this cell. Both PVM cells were somewhat swollen by the enlargement of internal vacuoles and membrane whorls. Large vacuoles lay close to the plasma membrane and sometimes appeared to be connected to whorls. Although most ER looked normal in both cells, some was somewhat swollen and more electron dense, suggesting that small vacuoles may occasionally derive directly from the ER. Each cell had a single axon extending toward the ventral nerve cord. This process contained several membrane whorls at the hillock, along its length, and at the growth cone. These axons were shorter than expected for this developmental stage (3 µm long), suggesting that outgrowth may have been disrupted.
Fig. 5. The early degeneration stage. A, PVM axon hillock in a glancing section. Many small membrane whorls are situated near the plasma membrane. B, A higher magnification view of a whorl from A, showing that it consists of onion skin-like concentric layers. C, The PVM soma in a second animal is swollen because of a large membrane-bound vacuole (vac), which may be in direct contact with plasma membrane. Arrows show small nested membrane whorls associated with the plasma membrane. ER, Endoplasmic reticulum; M, mitochondrion.
[View Larger Version of this Image (230K GIF file)]
Middle degeneration stage The two touch neurons (1 PVM, 1 AVM) at the middle degeneration stage were more swollen by vacuoles and whorls. Their nuclei were distorted and contained large clumps of chromatin. The ER and Golgi bodies were swollen, but the mitochondria were intact (Fig. 6). The cytoplasm of these neurons had fewer small whorls but was dominated, instead, by two to four very large vacuoles and/or whorls. The nucleus was displaced and deformed by invagination of a large vacuole in both cells (Fig. 6A,C). The cytoplasm was slightly less electron dense, suggesting that degradation of its contents had begun. No axons were seen in these cells. In addition, the AVM cell, but not the PVM neuron, was wrapped almost completely by a hypodermal process; only a small contact remained with the basal lamina (data not shown).
Fig. 6. The middle degeneration stage. A, The PVM nucleus (labeled with the cell name) is highly indented, invaded by a cytoplasmic vacuole (vac), and its chromatin is condensed. Several very large whorls of membrane cause cell swelling. The cytoplasm is slightly less electron-dense than that of nearby tissues. B, A different section of the same cell as in A, showing the mild swelling of the Golgi apparatus (G). C, A degenerating AVM cell has a huge vacuole (vac) invading the nucleus (labeled with the cell name), which is pushed to the edge of the soma. Cytoplasm of AVM is lighter than that of surrounding tissues. D, PVC soma in a late L1 stage (12 hr after hatching) deg-1(u38) larva is filled with large and small membrane whorls and vacuoles. The nucleus (not shown) was distorted in shape but otherwise normal in appearance. Magnification is the same in A-C.
[View Larger Version of this Image (164K GIF file)]
Late degeneration stage The appearance of the two late degeneration stage PVM neurons indicated a complete breakdown of cellular structures. One cell looked similar to the middle degeneration stage PVM cell described above, except for a local failure in the plasma membrane, which allowed some cytoplasmic contents to stream out from the cell (Fig. 7A). The nucleus was intact, but somewhat deformed, with very condensed chromatin. The cytoplasm was extremely light, as if degraded. The cytoplasm contained two very large vacuoles with associated whorled membranes that greatly distorted the cell. The cytoplasm also contained several intact mitochondria with moderate cisternal swelling, increased matrix density, and in one case a small outward bleb. Very few small whorls were seen. The second cell, which was large and distended into a lobular shape, had essentially no intact cytoplasmic contents, except for several compact dense bodies (Fig. 7B). No other signs of a nucleus remained, nor were any mitochondria present. One of these PVM cells (Fig. 7B) was surrounded completely by a hypodermal arm, which isolated the cell remnants from the basal lamina. The other PVM cell was not engulfed but remained in contact with the basal lamina.
Fig. 7. The late degeneration stage. A, PVM soma with a break in the plasma membrane in a ZB7 animal. Note the local damage to the closely apposed hypodermal tissue (H), where the cell contents of PVM have spilled out (open arrowhead). The PVM cytoplasm is very light, and the endoplasmic reticulum (ER) is swollen, but a mitochondrion (M) is still intact. The perimeter of the cell is irregular and is invaded by a finger of hypodermal tissue (xx). The soma is bordered by basal lamina on its right side (large arrowheads) that separates it from the intestine (INT). B, Another PVM soma has highly degraded cytoplasm, but the plasma membrane is intact. Several short projections extend from the swollen, distorted cell. The nucleus was not seen in serial thin sections of this cell. Small dense fragments (curved arrows) are each associated with a large membrane whorl. A thin hypodermal arm projects along the right side of the soma and separates it from the basal lamina (small arrowheads). In serial sections this arm completely engulfed the PVM cell.
[View Larger Version of this Image (195K GIF file)]
The pattern of neuronal death in deg-1(u38) animals, as seen in the degeneration of the PVC interneurons, was similar to that of the touch cells in mec-4 mutants. In an animal selected by light microscopy for having cells in the predegeneration stage, no abnormalities were found in any lumbar neurons, including the bilaterally symmetric PVCL and PVCR cells. In an animal with an early degeneration stage cell, the PVCR appeared normal, but the PVCL was moderately swollen with many large and small membrane whorls and vacuoles in the cytoplasm and a crenated nucleus with normal chromatin (Fig. 6D). In its pathology, PVCL looked intermediate between the early and middle stages of PVM degeneration in ZB7. Most organelles looked normal, although the ER was swollen near a few whorls and vacuoles. The electron density of the cytoplasm was normal. In an animal selected as having a middle degeneration death, the PVCL cell was normal, but the extremely swollen and lobulated PVCR neuron (data not shown) looked similar to a late-stage PVM death in ZB7. The cell had a highly distorted nucleus and a cytoplasm with very low electron density. Most cytoplasmic organelles were absent in PVCR; instead, the cell contained approximately one dozen large, loose membrane circles or whorls and much flocculent material that failed to embed well (in distinction to the surrounding cells that were well embedded). Overall, the progression of degeneration is strikingly similar in ZB7 and deg-1(u38) animals; the same sequence of cellular events is seen. Other signs of pathology in mec-4(gf) and deg-1(gf) animals
We examined other cells in wild-type, ZB7, and deg-1(u38) animals to determine the specificity of cell damage. Membrane whorls were rarely seen in wild-type animals. We found defects in three of nine animals; specifically, 3 of 120 body wall muscles, 1 of 9 anal depressor muscles, and 2 of 290 neuronal cell bodies had membrane whorls. In a survey of available prints of several hundred fixed animals with various genotypes, only one wild type, an animal incubated in colloidal gold before fixation, showed numerous whorls in muscles, neurons, and other cells in the head.
Cells next to the dying touch cells in ZB7 animals occasionally showed some pathology. In the vicinity of the ruptured late degeneration PVM cell, the hypodermal cytoplasm was damaged where the cytoplasmic contents of PVM had spilled (Fig. 7A).
Defects in neighboring cells also were seen when PVM cells had not lysed. For example, hypodermis, muscle, and neurons near the two early degeneration stage PVM cells showed minor local membrane infoldings or whorls that could be secondary responses to touch cell degeneration. A mild pathology (small membrane whorls and rare small vacuoles), which looked very similar to the earliest signs of toxicity in the touch neurons, was seen in cells that never contacted the touch cells in ZB7 animals. Although these defects may result from ectopic low-level expression of the mec-4(u231) transgene, some or all of this pathology may be attributable to treatment of the animals: 34 of 79 body wall muscles and 29 of 320 neurons in ZB7 animals mounted for light microscopy before fixation had whorls or vacuoles, whereas 4 of 75 body wall muscles and 8 of 200 neuronal cell bodies or axons in animals that were not mounted had them.
Mild pathology, usually membrane whorls and the occasional vacuole, also was seen in electron micrographs of several different cells in deg-1(u38) animals. These cells included neurons (e.g., some of the PHA chemosensory neurons; Fig. 8A), muscle cells, hypodermal cells, and other support cells. In the affected muscles, which included the anal depressor muscle (n = 2 of 3), dorsal body wall muscles (n = 5 of 13), and ventral body wall muscles (n = 3 of 12), membrane whorls were concentrated at muscle arms, muscle bellies, and distal extremities (Fig. 8B,C). Because these animals also were prepared for light microscopy before fixation, these ectopic signs of degeneration may have resulted from the handling of the animals. However, because empty vacuoles, which are not associated with cell deaths, have been seen by light microscopy throughout the animals (Chalfie and Wolinsky, 1990 
Fig. 8. Toxic changes in neurons and muscles in a deg-1(u38) larva. A, A membrane whorl and vacuole were found in a PHA dendrite in the phasmidial nerve. Similar whorls and vacuoles were found along the length of both PHA dendrites in this animal, but not in two other animals scored in serial thin sections. B, A membrane whorl is located at the boundary of the anal depressor muscle (AD) and a ventral body wall muscle (vm). C, Involution of the plasma membrane (curved arrow) in the anal depressor muscle (AD) has caused the insertion of membrane into the muscle cell nucleus (labeled with the cell name). The thin arrow indicates a smaller membrane whorl at the junction of the depressor muscle and a dorsal body wall muscle (dm).
[View Larger Version of this Image (177K GIF file)]
DISCUSSION
The progression of degenerative cell death
The degenerative death induced by gain-of-function alleles of the C. elegans degenerin genes mec-4 and deg-1 proceeds in a reproducible series of steps (summarized in Fig. 9). The earliest degenerative events are small infoldings of the plasma membrane. (More rarely, small swellings of the ER also occur.) In both mec-4 and deg-1 mutants the sites of the infoldings are distributed unevenly in the affected cells. In neurons the highest density of the infoldings was on the growing neurites, whereas in muscle cells infoldings were concentrated on the muscle bellies and muscle arms, areas specialized for cell-cell communication. We do not know the subcellular localization of the degenerin channels, but the infoldings could identify the positions of the channels. Alternatively, the high surface-to-volume ratio in these regions of the cell could render them more vulnerable to toxic insult.
Fig. 9. Stages in degenerin-induced cell death. Successive stages in PVM degeneration are depicted from the production of the cell-to-cell lysis. The first signs of pathology are the production of membrane infoldings and whorls. The cytoplasm becomes progressively less electron-dense as whorls and vacuoles accumulate and enlarge. These vacuoles deform the nucleus and dislocate it toward the plasma membrane. The axon seems to be disrupted by the middle degeneration stage. Mitochondria, Golgi, and endoplasmic reticulum, which are unaffected until late stages in degeneration, are not depicted.
[View Larger Version of this Image (28K GIF file)]
The number of active sites of infolding in the PVM cell increases from 2-5 per cell in the predegeneration stage to 15-22 in the early degeneration stage and then falls to 8-10 in the middle degeneration stage and none in the late degeneration stage. Because cells are dominated by a few very large whorls at later times, the infoldings may coalesce into the whorls with increasing numbers of layers. The whorls also may merge with each other, because neighboring whorls are sometimes surrounded by continuous outer shells. Although the layers of the whorls are packed tightly in early degeneration cells, they are packed more loosely as cell death progresses.
The proximity of early vacuoles to the infoldings at the cell surface suggests that vacuoles, like the whorls to which they are connected, originate from infoldings of plasma membrane. Because a few very large vacuoles are found in later stages, they may enlarge or coalesce from smaller vacuoles. If degenerin channels are concentrated in these membranes, as seems likely, the dramatic swelling seen in the dying cells could result from open channels creating osmotic differences between the internal and external compartments. The onion skin structure of the whorls would allow for the formation of an osmotic difference. Growing vacuoles and, perhaps, membrane whorls may contribute to the growing space next to the nucleus observed by light microscopy in early stage degenerations.
Although defects in the plasma and ER membranes are found early, alterations in nuclei and mitochondria occur late. Changes in the nuclei begin at the middle degeneration stage: chromatin progressively condenses as the nucleus invaginates, moves (or is displaced) toward the plasma membrane, and appears to fragment. In several cases, the electron microscopy data suggest that nuclear displacement results from the growth of a very large cytoplasmic vacuole. Mitochondria remain intact through early and middle stages, although occasional mitochondria show increased outer density at the earliest stages. Cristae show some swelling in middle and late-stage cells. Cytoplasmic degradation occurs gradually throughout the period of degeneration, perhaps by the activation of endogenous hydrolases (cf. Clarke, 1990
In the final stages of cell death organelles disappear, and only a few whorl remnants persist within the intact cell. We do not know whether, as seen with one late-stage PVM cell, the cell membrane usually ruptures as degeneration proceeds, but cell rupture could result in the sudden cell shrinkage that we saw by light microscopy in late-stage degenerations. The damage to surrounding cells seen in the ZB7 animals could be from the spillage of cell contents or from interactions with released membrane whorls. In any event, the dead cells eventually disappear, and serial thin sections of several older larvae show that no cellular debris remains (data not shown). This disappearance seems to require genes that also are needed for the engulfment of programmed cell deaths (Ellis et al., 1991 Degeneration pathology is dependent on degenerin gene dosage
Degenerin-induced cell death can be attributed to the expression of a toxic product. The dosage of the abnormal gene affects the onset of the cell death, e.g., the PVM and AVM deaths occur earlier in ZB7 animals than in mec-4(u231) mutants. Dosage of the toxic product also seems to influence the kinetics of degeneration. In ZB7 animals swollen touch receptor neurons disappear more quickly than they do in mec-4(u231) animals. Dosage also affects the onset of cell death of the PVC cell in deg-1(u38) mutants; cells die earlier in homozygous animals than in heterozygous animals (Chalfie and Wolinsky, 1990
mec-4 normally is expressed in the six touch receptor neurons, a restricted pattern of expression observed even when a mec-4lacZ fusion gene is present in multiple copies (Mitani et al., 1993 Similarities and differences with programmed cell death
Programmed cell death (Robertson and Thomson, 1982
Despite many differences, programmed cell death and degenerin-induced death share some features. Both cell deaths are characterized by the distortion of the nucleus, the condensation of chromatin (although condensation occurs less in the degenerin-induced deaths), the invagination of the initially spherical nucleus, and the eventual fragmentation of the nucleus. Both sets of dying cells are engulfed by the surrounding hypodermis. These similarities hint that some steps in both death processes might be shared. As far as the nuclear changes, however, any common mechanisms do not seem to require ced-3 or ced-4 function.
Similarities also have been seen between vertebrate apoptosis and necrosis: both types of cell death exhibit similar changes in nuclear morphology (chromatin clumping, nuclear fragmentation) (Wyllie, 1981 Inappropriate channel activity as a cause of cell pathology and death
Mutations near the putative pore-forming region or in an extracellular regulatory region of C. elegans degenerin proteins cause degenerative cell death (Driscoll and Chalfie, 1991subunit, encoded by the gene deg-3, results in a cell death that is morphologically similar to the degenerin-induced death (Treinin and Chalfie, 1995
This model for degenerin-induced death shares several features with that proposed for excitotoxic cell death as occurs in ischemia, hypoxia, epilepsy, and, possibly, neurodegenerative disease. In excitotoxic cell death, excess excitatory amino acids binding to glutamate receptors stimulate channel opening and result in excess Na+ and Ca2+ influx. This influx leads to the swelling and death of susceptible neurons (Choi, 1988
Increased ion channel open probability and osmotic imbalance can provoke endocytosis and vacuolation in other systems. For example, Mauthner cell giant synapses accumulate membrane cisternae or whorls and small vacuoles that follow increased endocytosis after direct electrical stimulation, which overwhelms homeostasis during recycling of synaptic vesicle membranes (Model et al., 1975
Defective ion channels underlie several autosomal dominant muscle diseases in mammals, including hyperkalemic periodic paralysis [HPP(+)] (caused by mutations in the SCNA sodium channel) (Ricker et al., 1994 )] (caused by mutations in CACNL1A3, the dihydropyridine receptor calcium channel) (Boerman et al., 1995
In weaver mutant mice a defective potassium channel (GIRK2) with altered gating and ion selectivity leads to selective cell death in the cerebellum, dentate gyrus, olfactory bulb, and germ line (Kofugi et al., 1995 A common theme in degenerative cell death?
A common feature of degenerin-induced death and the other conditions we have discussed is the production of membrane infoldings, membrane whorls, and vacuoles. These common features suggest that similar mechanisms may underlie several forms of cell pathology. We suggest that, in instances of membrane damage, the cell compensates by enhanced membrane cycling to dilute or to sequester harmful components localized to various membranes and to translocate them to degradative vacuoles. Such a surveillance mechanism may allow a variety of membranes to eliminate inappropriate, malfunctioning, or old components under normal conditions. In the cases of the degenerin-induced deaths, and perhaps other forms of neurodegeneration, this homeostatic mechanism is overwhelmed.
FOOTNOTES
Received July 9, 1996; revised Nov. 12, 1996; accepted Nov. 26, 1996.
This work was supported by National Institutes of Health Grant GM34775 to M.C. and National Science Foundation Grant IBN 92-0945 to M.D. We thank Jian Xue for determining the copy number of mec-4(u231) in ZB7, Eileen German for photography, and Laura Hall for help with Figure 9.
Correspondence should be addressed to Dr. Martin Chalfie, Department of Biological Sciences, 1012 Fairchild, Columbia University, 1212 Amsterdam Avenue, New York, NY 10027.
Dr. García-Añoveros' present address: Department of Neurobiology, Harvard Medical School, Massachusetts General Hospital, Wellman 414, Fruit Street, Boston, MA 02114.
REFERENCES
- Boerman RH, Ophoff RA, Links TP, van Eijk R, Sandkuijl LA, Elbaz A, Vale-Santos JE, Wintzen AR, van Deutekom JC, Isles DE, Fontaine B, Padberg GW, Frants RR (1995) Mutation in DHP receptor alpha 1 subunit (CACNL1A3) gene in a Dutch family with hypokalemic periodic paralysis. J Med Genet 32:44-47 .
[Abstract/Free Full Text] - Brenner S (1974) The genetics of Caenorhabditis elegans. Genetics 77:71-94 .
[Abstract/Free Full Text] - Canessa CM, Horisberger J-D, Rossier BC (1993) Epithelial sodium channel related to proteins involved in neurodegeneration. Nature 361:467-470 . [Medline]
- Canessa CM, Schild L, Buell G, Thorens B, Gautschi I, Horisberger J-D, Rossier BC (1994) Amiloride-sensitive epithelial Na+ channel is made of three homologous subunits. Nature 367:463-467 . [Medline]
- Cannon SC, Hayward LJ, Beech J, Brown Jr RH (1995) Sodium channel inactivation is impaired in equine hyperkalemic periodic paralysis. J Neurophysiol 73:1892-1899 .
[Abstract/Free Full Text] - Chalfie M, Au M (1989) Genetic control of differentiation of the Caenorhabditis elegans touch receptor neurons. Science 243:1027-1033 .
[Abstract/Free Full Text] - Chalfie M, Sulston J (1981) Developmental genetics of the mechanosensory neurons of Caenorhabditis elegans. Dev Biol 82:358-370 . [Web of Science][Medline]
- Chalfie M, Wolinsky E (1990) The identification and suppression of inherited neurodegeneration in Caenorhabditis elegans. Nature 345:410-416 . [Medline]
- Chalfie M, Driscoll M, Huang M (1993) Degenerin similarities. Nature 361:504 . [Medline]
- Choi DW (1988) Glutamate neurotoxicity and diseases of the nervous system. Neuron 1:623-634 . [Web of Science][Medline]
- Chung SK, Cohen RS, Pfaff DW (1990) Transneuronal degeneration in the midbrain central gray following chemical lesions in the ventromedial nucleus: a qualitative and quantitative analysis. Neuroscience 38:409-426 . [Web of Science][Medline]
- Clarke PGH (1990) Developmental cell death: morphological diversity and multiple mechanisms. Anat Embryol (Berl) 181:195-213. [Medline]
- Driscoll M, Chalfie M (1991) The mec-4 gene is a member of a family of Caenorhabditis elegans genes that can mutate to induce neuronal degeneration. Nature 349:588-593 . [Medline]
- Elbaz A, Vale-Santos J, Jurkatt-Rott K, Lapie P, Ophoff RA, Bady B, Links TP, Piussan C, Vila A, Monnier N, Padberg GW, Abe K, Feingold N, Guimares J, Wintzen AR, van der Hoeven JH, Saudubray JM, Grunfeld JP, Lenoir G, Nivet H, Echenne B, Frants RR, Fardeau M, Lehmann-Horn F, Fontaine B (1995) Hypokalemic periodic paralysis and the dihydropyridine receptor (CACNL1A3): genotype/phenotype correlations for two predominant mutations and evidence for the absence of a founder effect in 16 Caucasian families. Am J Hum Genet 56:374-380 . [Web of Science][Medline]
- Ellis H, Horvitz HR (1986) Genetic control of programmed cell death in the nematode C. elegans. Cell 44:817-829 . [Web of Science][Medline]
- Ellis RE, Yuan JY, Horvitz HR (1991) Mechanisms and functions of cell death. Annu Rev Cell Biol 7:663-698 . [Web of Science]
- Engel AG (1970) Evolution and content of vacuoles in primary hypokalemic periodic paralysis. Mayo Clin Proc 45:774-814 . [Web of Science][Medline]
- Fejtl M, Szarowski DH, Decker D, Buttle K, Carpenter DO, Turner JN (1995) Three-dimensional imaging and electrophysiology of live Aplysia neurons during volume perturbation: confocal light and high-voltage electron microscopy. J Microsc Soc Am 1:75-85.
- Fernandez PA, Potello R, Rangini Z, Doupe A, Drexler HC, Yuan J (1994) Expression of a specific marker of avian programmed cell death in both apoptosis and necrosis. Proc Natl Acad Sci USA 91:8641-8645 .
[Abstract/Free Full Text] - Fire A (1986) Integrative transformation of Caenorhabditis elegans. EMBO J 5:2673-2680. [Web of Science][Medline]
- Fire A, Harrison SW, Dixon D (1990) A modular set of lacZ fusion vectors for studying gene expression in C. elegans. Gene 93:189-198 . [Web of Science][Medline]
- García-Añoveros J (1995) Genetic analysis of degenerin proteins in Caenorhabditis elegans. PhD thesis, Columbia University, New York.
- García-Añoveros J, Ma C, Chalfie M (1995) Regulation of Caenorhabditis elegans degenerin proteins by a putative extracellular domain. Curr Biol 5:441-448. [Medline]
- Hajos F, Garthwaite G, Garthwaite JG (1986) Reversible and irreversible neuronal damage caused by excitatory amino acid analogues in rat cerebellar slices. Neuroscience 18:417-436 . [Web of Science][Medline]
- Hall DH (1995) Electron microscopy and 3D image reconstruction. In: C. elegans: modern biological analysis of an organism, Vol 48, Methods in cell biology (Epstein HF, Shakes DC, eds), pp 395-436. San Diego: Academic.
- Hedgecock EM, Sulston JE, Thomson JN (1983) Mutations affecting programmed cell deaths in the nematode Caenorhabditis elegans. Science 220:1277-1279 .
[Abstract/Free Full Text] - Hengartner M, Horvitz HR (1994) C. elegans survival gene ced-9 encodes a functional homolog of the mammalian proto-oncogene bcl-2. Cell 76:665-676 . [Web of Science][Medline]
- Herman RK (1987) Mosaic analysis of two genes that affect nervous system structure in Caenorhabditis elegans. Genetics 116:377-388 .
[Abstract/Free Full Text] - Hong K, Driscoll M (1994) A transmembrane domain of the putative channel subunit MEC-4 influences mechanotransduction and neurodegeneration in C. elegans. Nature 367:470-473 . [Medline]
- Huang M, Chalfie M (1994) Gene interactions affecting mechanosensory transduction in Caenorhabditis elegans. Nature 367:467-470 . [Medline]
- Hudson AJ, Ebers GC, Bulman DE (1995) The skeletal muscle sodium and chloride channel diseases. Brain 118:547-563 .
[Abstract/Free Full Text] - Kramer JM, Johnson JJ (1993) Analysis of mutations in the sqt-1 and rol-6 collagen genes of Caenorhabditis elegans. Genetics 135:1035-1045 . [Abstract]
- Kofugi P, Davidson N, Lester HA (1995) Evidence that neuronal G-protein-gated inwardly rectifying K+ channels are activated by G
subunits and function as heterodimers. Proc Natl Acad Sci USA 92:6542-6546.
[Abstract/Free Full Text] - Kofugi P, Hofer M, Millen KJ, Davidson N, Lester HA, Hatten ME (1996) Functional analysis of the weaver mutant GIRK2 K+ channel and rescue of weaver granule cells. Neuron 16:941-952. [Web of Science][Medline]
- Manoil C (1990) Analysis of protein localization by use of gene fusion with complementary properties. J Bacteriol 172:1035-1042 .
[Abstract/Free Full Text] - Martin DP, Schmidt RE, DiStefano PS, Lowry OH, Carter JG, Johnson Jr EM (1988) Inhibitors of protein synthesis prevent neuronal death caused by nerve growth factor deprivation. J Cell Biol 106:829-844 .
[Abstract/Free Full Text] - Mello CC, Kramer JM, Stinchcomb D, Ambros V (1991) Efficient gene transfer in C. elegans: extrachromosomal maintenance and integration of transforming sequences. EMBO J 10:3959-3970 . [Web of Science][Medline]
- Mitani S, Du H, Hall DH, Driscoll M, Chalfie M (1993) Combinatorial control of touch neuron receptor expression in Caenorhabditis elegans. Development (Camb) 119:773-783 . [Abstract]
- Miura M, Zhu H, Rotello R, Hartweig EA, Yuan J (1993) Induction of apoptosis in fibroblasts by IL-1
- Model PG, Highstein SM, Bennett MVL (1975) Depletion of vesicles and fatigue of transmission at a vertebrate central synapse. Brain Res 98:209-228 . [Web of Science][Medline]
- Olney JW (1971) Glutamate-induced neuronal necrosis in the infant mouse hypothalamus. An electron microscopic study. J Neuropathol Exp Neurol 30:75-90 . [Web of Science][Medline]
- Olney JW, Rhee V, Ho OL (1974) Kainic acid: a powerful neurotoxic analogue of glutamate. Brain Res 77:507-512 . [Web of Science][Medline]
- Patil N, Cox DR, Bhat D, Faham M, Myers RM, Peterson AS (1995) A potassium channel mutation in weaver mice implicates membrane excitability in granule cell differentiation. Nat Genet 11:126-129 . [Web of Science][Medline]
- Rakic P, Sidman RL (1973a) Sequence of developmental abnormalities leading to granule cell deficit in cerebellar cortex of weaver mutant mice. J Comp Neurol 152:103-132 . [Web of Science][Medline]
- Rakic P, Sidman RL (1973b) weaver mutant mouse cerebellum: defective neuronal migration secondary to specific abnormality of Bergmann glia. Proc Natl Acad Sci USA 70:240-244 .
[Abstract/Free Full Text] - Ricker K, Moxley III RT, Heine R, Lehmann-Horn F (1994) Myotonia fluctuans: a third type of muscle sodium channel disease. Arch Neurol 51:1095-1102 .
[Abstract/Free Full Text] - Robertson AMG, Thomson JN (1982) Morphology of programmed cell death in the ventral nerve cord of Caenorhabditis elegans larvae. J Embryol Exp Morphol 67:89-100. [Web of Science]
- Rothman SM (1994) Excitotoxic neuronal death: mechanisms and clinical relevance. Semin Neurosci 6:315-322.
- Salahuddin TS, Johansson BB, Kalimo H, Olsson Y (1988) Structural changes in the rat brain after carotid infusions of hyperosmolar solutions. Acta Neuropathol (Berl) 77:5-13 . [Medline]
- Sansone V, Rotondo G, Ptacek LJ, Meola G (1994) Mutation in the S4 segment of the adult skeletal sodium channel gene in an Italian paramyotonia congenita (PC) family. Ital J Neurol Sci 15:473-480 . [Web of Science][Medline]
- Shreffler W, Margardino T, Shekdar C, Wolinsky E (1995) The unc-8 and sup-40 genes regulate ion channel function in Caenorhabditis elegans motor neurons. Genetics 139:1261-1272 . [Abstract]
- Slesinger PA, Patil N, Liao J, Jan YN, Jan LY, Cox DR (1996) Functional effects of the mouse weaver mutation on G-protein-gated inwardly rectifying K+ channels. Neuron 16:321-331 . [Web of Science][Medline]
- Sperk G, Lassmann H, Baran H, Kish SJ, Seitelberger H, Hornytiewícz O (1983) Kainic acid-induced seizures: neurochemical and histopathological changes. Neuroscience 10:1301-1315 . [Web of Science][Medline]
- Spier SJ, Carlson GP, Holliday TA, Cardinet III GH, Pickar JG (1990) Hyperkalemic periodic paralysis in horses. J Am Vet Med Assoc 197:1009-1017 . [Web of Science][Medline]
- Sulston JE, Horvitz HR (1977) Post-embryonic cell lineages of the nematode Caenorhabditis elegans. Dev Biol 56:110-156 . [Web of Science][Medline]
- Treinin M, Chalfie M (1995) A mutated acetylcholine receptor subunit causes neuronal degeneration in C. elegans. Neuron 14:871-877 . [Web of Science][Medline]
- Trump BF, Berezesky IK (1985) Cellular ion regulation and disease: a hypothesis. Curr Top Membr Transp 25:279-319.
- Vaux DL, Weissman IL, Kim SK (1992) Prevention of programmed cell death in Caenorhabditis elegans by human bcl-2. Science 258:1955-1957 .
[Abstract/Free Full Text] - Verina T, Tang X, Fitzpatrick L, Norton J, Vogelweid C, Ghetti B (1995) Degeneration of Sertoli and spermatogenic cells in homozygous and heterozygous weaver mice. J Neurogenet 9:251-265 . [Web of Science][Medline]
- Waldmann R, Champigny G, Bassilana F, Voilley N, Lazdunski M (1995) Molecular cloning and functional expression of a novel amiloride-sensitive Na+ channel. J Biol Chem 270:27411-27414 .
[Abstract/Free Full Text] - Waldmann R, Champigny G, Voilley N, Lauritzen I, Lazdunski M (1996) The mammalian degenerin MDEG, an amiloride-sensitive cation channel activated by mutations causing degeneration in Caenorhabditis elegans. J Biol Chem 271:10433-10436 .
[Abstract/Free Full Text] - Wasterlain CG, Shirasaka Y (1994) Seizures, brain damage, and brain development. Brain Dev 16:279-295 . [Web of Science][Medline]
- White JG, Southgate E, Thomson JN, Brenner S (1986) The structure of the nervous system of Caenorhabditis elegans. Philos Trans R Soc Lond [Biol] 314:1-340.
- Wyllie AH (1981) Cell death: a new classification separating apoptosis from necrosis. In: Cell death in biology and pathology (Bowen ID, Lockshin RA, eds), pp 9-34. New York: Chapman and Hall.
- Wyllie AH, Kerr JFR, Currie AR (1980) Cell death: the significance of apoptosis. Int Rev Cytol 68:251-306 . [Medline]
- Yuan JY, Horvitz HR (1990) The Caenorhabditis elegans genes ced-3 and ced-4 act cell autonomously to cause programmed cell death. Dev Biol 138:33-41 . [Web of Science][Medline]
- Yuan J, Shaham S, Ledoux S, Ellis HM, Horvitz HR (1993) The C. elegans cell death gene ced-3 encodes a protein similar to mammalian interleukin-1
- Zhou J, Spier SJ, Beech J, Hoffman EP (1994) Pathophysiology of sodium channelopathies: correlation of normal/mutant mRNA ratios with clinical phenotype in dominantly inherited periodic paralysis. Hum Mol Genet 3:1599-1603 .
[Abstract/Free Full Text]
Copyright ©1997 Society for Neuroscience 0270-6474/1997/171033-13$05.00/0
This article has been cited by other articles:
B. D. Galvin, S. Kim, and H. R. Horvitz
Caenorhabditis elegans Genes Required for the Engulfment of Apoptotic Corpses Function in the Cytotoxic Cell Deaths Induced by Mutations in lin-24 and lin-33
Genetics, May 1, 2008; 179(1): 403 - 417.
[Abstract] [Full Text] [PDF]
M. L. Toth, P. Simon, A. L. Kovacs, and T. Vellai
Influence of autophagy genes on ion-channel-dependent neuronal degeneration in Caenorhabditis elegans
J. Cell Sci., March 15, 2007; 120(6): 1134 - 1141.
[Abstract] [Full Text] [PDF]
T. Takasaki, Z. Liu, Y. Habara, K. Nishiwaki, J.-i. Nakayama, K. Inoue, H. Sakamoto, and S. Strome
MRG-1, an autosome-associated protein, silences X-linked genes and protects germline immortality in Caenorhabditis elegans
Development, February 15, 2007; 134(4): 757 - 767.
[Abstract] [Full Text] [PDF]
M. Artal-Sanz, C. Samara, P. Syntichaki, and N. Tavernarakis
Lysosomal biogenesis and function is critical for necrotic cell death in Caenorhabditis elegans
J. Cell Biol., April 24, 2006; 173(2): 231 - 239.
[Abstract] [Full Text] [PDF]
D. C. Royal, L. Bianchi, M. A. Royal, M. Lizzio Jr., G. Mukherjee, Y. O. Nunez, and M. Driscoll
Temperature-sensitive Mutant of the Caenorhabditis elegans Neurotoxic MEC-4(d) DEG/ENaC Channel Identifies a Site Required for Trafficking or Surface Maintenance
J. Biol. Chem., December 23, 2005; 280(51): 41976 - 41986.
[Abstract] [Full Text] [PDF]
A. Kosta, C. Roisin-Bouffay, M.-F. Luciani, G. P. Otto, R. H. Kessin, and P. Golstein
Autophagy Gene Disruption Reveals a Non-vacuolar Cell Death Pathway in Dictyostelium
J. Biol. Chem., November 12, 2004; 279(46): 48404 - 48409.
[Abstract] [Full Text] [PDF]
M. T. Crow, K. Mani, Y.-J. Nam, and R. N. Kitsis
The Mitochondrial Death Pathway and Cardiac Myocyte Apoptosis
Circ. Res., November 12, 2004; 95(10): 957 - 970.
[Abstract] [Full Text] [PDF]
P. Syntichaki and N. Tavernarakis
Genetic Models of Mechanotransduction: The Nematode Caenorhabditis elegans
Physiol Rev, October 1, 2004; 84(4): 1097 - 1153.
[Abstract] [Full Text] [PDF]
B. C. Kraemer, B. Zhang, J. B. Leverenz, J. H. Thomas, J. Q. Trojanowski, and G. D. Schellenberg
From the Cover: Neurodegeneration and defective neurotransmission in a Caenorhabditis elegans model of tauopathy
PNAS, August 19, 2003; 100(17): 9980 - 9985.
[Abstract] [Full Text] [PDF]
A. S. Toker, Y. Teng, H. B. Ferreira, S. W. Emmons, and M. Chalfie
The Caenorhabditis elegans spalt-like gene sem-4 restricts touch cell fate by repressing the selector Hox gene egl-5 and the effector gene mec-3
Development, August 15, 2003; 130(16): 3831 - 3840.
[Abstract] [Full Text] [PDF]
K. Strange
From Genes to Integrative Physiology: Ion Channel and Transporter Biology in Caenorhabditis elegans
Physiol Rev, April 1, 2003; 83(2): 377 - 415.
[Abstract] [Full Text] [PDF]
S. Kellenberger and L. Schild
Epithelial Sodium Channel/Degenerin Family of Ion Channels: A Variety of Functions for a Shared Structure
Physiol Rev, July 1, 2002; 82(3): 735 - 767.
[Abstract] [Full Text] [PDF]
M. J. Palladino, T. J. Hadley, and B. Ganetzky
Temperature-Sensitive Paralytic Mutants Are Enriched For Those Causing Neurodegeneration in Drosophila
Genetics, July 1, 2002; 161(3): 1197 - 1208.
[Abstract] [Full Text] [PDF]
P. J. Muhlrad and S. Ward
Spermiogenesis Initiation in Caenorhabditis elegans Involves a Casein Kinase 1 Encoded by the spe-6 Gene
Genetics, May 1, 2002; 161(1): 143 - 155.
[Abstract] [Full Text] [PDF]
R. Nass, D. H. Hall, D. M. Miller III, and R. D. Blakely
Neurotoxin-induced degeneration of dopamine neurons in Caenorhabditis elegans
PNAS, February 20, 2002; (2002) 42497999.
[Abstract] [Full Text] [PDF]
K. R. Svoboda, A. E. Linares, and A. B. Ribera
Activity regulates programmed cell death of zebrafish Rohon-Beard neurons
Development, September 15, 2001; 128(18): 3511 - 3520.
[Abstract] [Full Text] [PDF]
A. H. Wyllie and P. Golstein
More than one way to go
PNAS, January 2, 2001; 98(1): 11 - 13.
[Full Text] [PDF]
J. Hewett, C. Gonzalez-Agosti, D. Slater, P. Ziefer, S. Li, D. Bergeron, D. J. Jacoby, L. J. Ozelius, V. Ramesh, and X. O. Breakefield
Mutant torsinA, responsible for early-onset torsion dystonia, forms membrane inclusions in cultured neural cells
Hum. Mol. Genet., May 22, 2000; 9(9): 1403 - 1413.
[Abstract] [Full Text] [PDF]
H. Sakai, E. Lingueglia, G. Champigny, M.-G. Mattei, and M. Lazdunski
Cloning and functional expression of a novel degenerin-like Na+ channel gene in mammals
J. Physiol., September 1, 1999; 519(2): 323 - 333.
[Abstract] [Full Text] [PDF]
S Kim, X. Ren, E Fox, and W. Wadsworth
SDQR migrations in Caenorhabditis elegans are controlled by multiple guidance cues and changing responses to netrin UNC-6
Development, January 9, 1999; 126(17): 3881 - 3890.
[Abstract] [PDF]
The Phenotype of mes-2, mes-3, mes-4 and mes-6, Maternal-Effect Genes Required for Survival of the Germline in Caenorhabditis elegans, Is Sensitive to Chromosome Dosage
Genetics, January 1, 1998; 148(1): 167 - 186.
E. Lingueglia, Jan. R. de Weille, F. Bassilana, C. Heurteaux, H. Sakai, R. Waldmann, and M. Lazdunski
A Modulatory Subunit of Acid Sensing Ion Channels in Brain and Dorsal Root Ganglion Cells
J. Biol. Chem., November 21, 1997; 272(47): 29778 - 29783.
[Abstract] [Full Text] [PDF]
R. Nass, D. H. Hall, D. M. Miller III, and R. D. Blakely
Neurotoxin-induced degeneration of dopamine neurons in Caenorhabditis elegans
PNAS, March 5, 2002; 99(5): 3264 - 3269.
[Abstract] [Full Text] [PDF]
This Article Abstract 
Full Text (PDF) Submit an eLetter Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager 
Citing Articles Citing Articles via HighWire Citing Articles via Web of Science (79) Citing Articles via Google Scholar Google Scholar Articles by Hall, D. H. Articles by Driscoll, M. Search for Related Content PubMed PubMed Citation Articles by Hall, D. H. Articles by Driscoll, M.
ABSTRACT
|
|
|
|
 |
|