Distinct susceptibility of developing neurons to death following Bax overexpression in the chicken embryo.

Bax is a proapoptotic protein that is required for programmed cell death (PCD) of many neuronal populations. Here we show that, during an early period of retinal PCD and in naturally occurring sensory and motor neuron (MN) death in the spinal cord, Bax delivery results in enhanced death of these neural populations. In contrast, Bax overexpression fails to enhance an early phase of MN death that occurs in the cervical spinal cord, although overexpressed Bax appears to be activated in dying MNs. Bax overexpression does not also affect the survival of immature neurons prior to the PCD period. Taken together, these data provide the first in vivo evidence suggesting that Bax appears to act selectively as an executioner only in neurons undergoing PCD. Furthermore, although Bax appears to mediate the execution pathway for PCD, the effect of Bax overexpression on susceptibility to death differs between different neuronal populations.

Low temperature stimulates alpha-melanophore-stimulating hormone secretion and inhibits background adaptation in Xenopus laevis.

It is well-known that alpha-melanophore-stimulating hormone (alpha-MSH) release from the amphibian pars intermedia (PI) depends on the light condition of the animal's background, permitting the animal to adapt the colour of its skin to background light intensity. In the present study, we carried out nine experiments on the effect of low temperature on this skin adaptation process in the toad Xenopus laevis, using the skin melanophore index (MI) bioassay and a radioimmunoassay to measure skin colour adaptation and alpha-MSH secretion, respectively. We show that temperatures below 8 degrees C stimulate alpha-MSH secretion and skin darkening, with a maximum at 5 degrees C, independent of the illumination state of the background. No significant stimulatory effect of low temperature on the MI and alpha-MSH plasma contents was noted when the experiment was repeated with toads from which the neurointermediate lobe (NIL) had been surgically extirpated. This indicates that low temperature stimulates alpha-MSH release from melanotrope cells located in the PI. An in vitro superfusion study with the NIL demonstrated that low temperature does not act directly on the PI. A possible role of the central nervous system in cold-induced alpha-MSH release from the PI was tested by studying the hypothalamic expression of c-Fos (as an indicator for neuronal activity) and the coexistence of c-Fos with the regulators of melanotrope cell activity, neuropeptide Y (NPY) and thyrotrophin-releasing hormone (TRH), using double fluorescence immunocytochemistry. Upon lowering temperature from 22 degrees C to 5 degrees C, in white-adapted animals c-Fos expression decreased in NPY-producing suprachiasmatic-melanotrope-inhibiting neurones (SMIN) in the ventrolateral area of the suprachiasmatic nucleus (SC) but increased in TRH-containing neurones of the magnocellular nucleus. TRH is known to stimulate melanotrope alpha-MSH release. We conclude that temperatures around 5 degrees C inactivate the SMIN in the SC and activate TRH-neurones in the magnocellular nucleus, resulting in enhanced alpha-MSH secretion from the PI, darkening the skin of white-adapted X. laevis.

CrmA gene expression protects mice against concanavalin-A-induced hepatitis by inhibiting IL-18 secretion and hepatocyte apoptosis.

Activated cytotoxic T-cell-mediated hepatocyte apoptosis via Fas/Fas-ligand and perforin/granzyme pathways are believed to involve the model of concanavalin A (ConA)-induced hepatitis. The purpose of the present study is to investigate whether the cytokine response modifier A (crmA) gene effectively inhibits the hepatocyte apoptosis of ConA-induced hepatitis. We examined survival rates, liver pathology, immune histological changes, and cytokine profiles from mice receiving the recombinant adenovirus vectors containing cre and/or crmA genes, transferred to the liver 3 days before ConA injection, and a crmA gene nonexpression control group. Injection of ConA into mice rapidly led to massive hepatocyte apoptosis, and infiltration of leukocytes, especially CD11b(+) inflammatory cells. In contrast, liver damage was dramatically reduced in the mice that expressed the crmA gene. However, infiltration by CD4(+) cells was not affected. The survival of the mice increased significantly to 100% in the treated group versus the control group. Furthermore, we demonstrated that interleukin (IL)-18 plays an important role in ConA-induced hepatitis, and that crmA expression significantly inhibited IL-18 secretion. Our results showed that the crmA gene effectively inhibits apoptosis induced by ConA hepatitis. This indicates a potential therapeutic usage of crmA for protection from cellular damage due to hepatitis.

Caspase activity is involved in, but is dispensable for, early motoneuron death in the chick embryo cervical spinal cord.

We examined the role of caspases in the early programmed cell death (PCD) of motoneurons (MNs) in the chick embryo cervical cord between embryonic day (E) 4 and E5. An increase in caspase-3-like activity in MNs was observed at E4.5. Treatment with an inhibitor of caspase-3-like activity, Ac-DEVD-CHO, for 12 h blocked this increase and revealed that caspase-3-like activity is mainly responsible for DNA fragmentation and the nuclear changes during PCD but not for degenerative changes in the cytoplasm. When a more broad-spectrum caspase inhibitor was used (bocaspartyl (OMe)-fluoromethyl ketone, BAF), the appearance of degenerative changes in the cytoplasm was delayed by at least 12 h. However, following treatment with either Ac-DEVD-CHO or BAF for 24 h, the number of surviving healthy MNs did not differ from controls, indicating a normal occurrence of PCD despite the inhibition of caspases. These results suggest that caspase cascades that occur upstream of and are independent of the activation of caspase-3-like activity are responsible for the degenerative changes in the cytoplasm of dying cervical MNs. These data also suggest that, although one function of caspases may be to facilitate the kinetics of PCD, caspases are nonetheless dispensable for at least some forms of normal neuronal PCD in vivo.

In vitro prevention of cell-mediated xeno-graft rejection via the Fas/FasL-pathway in CrmA-transducted porcine kidney cells.

Cell-mediated cytotoxicity may be involved in delayed and/or chronic xenograft rejection in which apoptosis is induced in the grafted cells via the Fas/Fas-ligand (FasL) and perforin/granzyme pathways. One barrier to the potential use of xeonogenic grafts for humans may be Fas/FasL-mediated apoptosis, which would be blocked by the gene expression of cytokine response modifier A (CrmA), a cowpox virus gene product. The purpose of this study is to explore whether crmA is an effective candidate gene for inhibiting apoptosis in an in vitro model of xenograft rejection, using Fas-expressing non-primate cells cultured with a soluble recombinant human FasL (sFasL). A recombinant adenovirus vector expressing CrmA (AxCALNLCrmA) was successfully generated with a Cre-mediated switching system. PK15 cells, derived from a porcine kidney and infected with AxCALNLCrmA and/or AxCANCre at a multiplicity of infection (MOI) ranging from 0.1 to 100, were cultured with human sFasL derived from KFL74.18, a human FasL-overexpressed cell line. The gene-expression level of the PK15 cells was confirmed by CrmA-immune staining. Approximately 70% of the control PK15 cells showed induced apoptosis when cultured with sFasL. In contrast, the apoptosis was dramatically reduced in crmA-gene-transduced PK15 cells. The inhibitory effect of apoptosis increased with an increase in the infection dose of AxCANCre. In addition, the activity of caspases 3 and 8 was significantly inhibited in the crmA-transduced cells. These results indicate that CrmA is an effective gene product for inhibiting Fas/FasL-mediated apoptosis, which suggests the potential therapeutic use of its gene transduction to protect against graft damage due to delayed and/or chronic xenograft rejection.

Comparison of sensory and sympathetic innervation of the dura mater and posterior longitudinal ligament in the cervical spine after removal of the stellate ganglion.

Although cervical spinal tissues are rich in sensory and sympathetic fibers, which play a significant role in clinical phenomena, there is little information available regarding their anatomical characteristics. In this study, we compared the innervation of the cervical dura mater and the posterior longitudinal ligament (PLL) to that after removal of the stellate ganglion to determine whether the anatomical background plays a significant role in clinical manifestations. Immunoreactivities for calcitonin gene-related peptide (CGRP) and substance P (SP) were used as sensory markers, and immunoreactivity for neuropeptide Y (NPY) was used as a sympathetic marker. Sensory fibers in the cervical dura mater were distributed within each cervical segment, but those in the PLL extended beyond the segmental borders. A dense sensory fiber network forming a single layer was seen at the intervertebral disc region in the cervical PLL, whereas sympathetic fibers in this region were sparsely distributed. Sympathetic fibers were distributed not only around the vascular wall but also in the region independent from vessels, and some occasionally ran together with sensory fibers in both the dura mater and the PLL. Removal of the stellate ganglion had little effect on the distribution of sensory fibers but denervated the sympathetic fiber networks in the region independent from vessels of the upper ipsilateral cervical PLL. In conclusion, the cervical dura mater and the PLL have different sensory and sympathetic innervations. Sympathetic fibers pass through the stellate ganglion to project to the region independent from vessels in the upper cervical PLL. Clinical symptoms may be attributed to this characteristic innervation of the cervical spine.

Roles of caspases in the programmed cell death of motoneurons in vivo.

Cysteine proteases comprising the caspase family have been considered one of the major executioners of programmed cell death. However, detailed analyses of the programmed cell death of developing motoneurons in mice following the genetic deletion of two key caspases, casp-3 and casp-9, and in the chick embryo following treatment with caspase inhibitors, indicate that normal amounts of cell loss occur although the death process is delayed. Motoneurons undergoing programmed cell death without caspase activities exhibit a nonapoptotic morphology in which nuclear changes such as chromatin condensation are absent or reduced and which exhibit extensive cytoplasmic vacuolization such as is rarely observed in degenerating control neurons. These results suggest that caspases are involved in, but are not indispensable for, the developmental death of motoneurons, and that one function of caspases may be to facilitate the removal of cells that are destined to die. Possible alternative caspase-independent pathways for the programmed death of motoneurons are discussed.

Inhibition of Fas-mediated fulminant hepatitis in CrmA gene-transfected mice.

Hyperimmune response via Fas/Fas-ligand and perforin/granzyme pathways may be essential in pathogenesis of virus-induced fulminant hepatitis. CrmA inhibits activation of caspases and granzyme B, suggesting it may block these pathways. We investigated whether CrmA expression would inhibit Fas-associated lethal hepatitis in mice. We successfully generated AxCALNLCrmA, a recombinant adenovirus expressing CrmA gene with a Cre-mediated switching cassette. We increased CrmA expression level in the liver transfected with AxCALNLCrmA (10(9) pfu) by increasing administration dose (10(7)-10(9) pfu) of AxCANCre, a recombinant, adenovirus-expressing Cre gene. Injection of anti-Fas antibody into the control mice rapidly led to animal death due to massive liver apoptosis, while the apoptosis was dramatically reduced in the CrmA-expressed mice. The animal survival increased with an increase of CrmA expression. The formation of active caspase-3 was markedly inhibited in the crmA-transfected hepatocytes in vitro. These results suggest that crmA is an effective gene that can inhibit immune-related liver apoptosis.

Expression pattern of GDNF, c-ret, and GFRalphas suggests novel roles for GDNF ligands during early organogenesis in the chick embryo.

We have cloned a partial cDNA of chicken glial cell line-derived neurotrophic factor (GDNF) and systematically examined its expression pattern as well as that of GDNF-binding components (GDNF family receptor alpha-1 and 2: GFRalpha-1 and 2) and a common signal transduction receptor (c-ret protooncogene: RET) during very early developmental stages. In addition, we also examined the expression pattern of an apparent avian-specific binding component, GFRalpha-4. The cloned chicken cDNA for GDNF had approximately 80% homology to mammalian counterparts. The expression of GDNF mRNA occurred in many spatially and temporally discrete regions such as the intermediate mesoderm, the floor plate of the spinal cord, pharyngeal endoderm contacting the epibranchial placodes, distal ganglia of cranial nerves, subpopulations of mesenchyme cells in the craniofacial region, and in the mesodermal wall of the digestive tract. Both a GDNF receptor signal transduction component (RET) and a binding component (GFRalpha-1 or GFRalpha-2) were independently expressed in nearby interacting tissues such as the somites, peripheral and central nervous system, and mesenchyme cells in the craniofacial region. These observations suggest that possible combinations of novel unidentified receptors acting with RET or with GFRalphas may mediate GDNF-derived signals and indicate that GDNF or other family members may have previously unidentified actions in early organogenesis in the chick embryo.

Suppressive effects of Neiting acupuncture on toothache: an experimental analysis on Fos expression evoked by tooth pulp stimulation in the trigeminal subnucleus pars caudalis and the periaqueductal gray of rats.

To clarify the antinociceptive mechanism of acupuncture on acute pain, c-fos protein (Fos) expression induced by tooth pulp stimulation was immunohistochemically examined in the spinal trigeminal subnucleus pars caudalis (spVc) and the periaqueductal gray (PAG) of rats with or without Neiting acupuncture. The central projection of trigeminal ganglion neurons innervating in the tooth pulp was examined by tract-tracing method with horseradish peroxidase-conjugated wheat germ agglutinin (WGA-HRP). Central terminals from the first maxillary molar tooth were labeled transganglionically in the dorsomedial part of spVc with WGA-HRP. Numerous numbers of Fos-immunoreactive (Fos-ir) cells were found in the spVc and PAG by stimulation of the tooth pulp with acetic acid or saline. Neiting acupuncture significantly reduced the Fos expression in the spVc induced by tooth pulp stimulation. On the other hand, Neiting acupuncture evoked many Fos-ir cells in the PAG. The present results suggest that Neiting acupuncture activated PAG neurons that sent descending inhibitory fibers to medullo-spinal nociceptive neurons, and reduced the number of Fos-expressed neurons in the trigeminal subnucleus pars caudalis mediating noxious information from teeth to the higher central nervous system.

Possible participation of intracellular platelet-activating factor in tumor necrosis factor-alpha production by rat peritoneal macrophages.

Stimulation of rat peritoneal macrophages by thapsigargin (46.1 nM) increased levels of tumor necrosis factor-alpha and prostaglandin E2 in the conditioned medium. Platelet-activating factor (PAF) was not detected in the conditioned medium, but the level of cell-associated PAF was increased transiently by thapsigargin. The PAF receptor antagonists such as E6123 ((S)-(+)-6-(2-chlorophenyl)-3-cyclopro-panecarbonyl-8,11-dim ethyl-2,3,4,5-tetrahydro-8 H-pyrido[4',3':4,5]thieno [3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepine), L-652,73 1 (2,5-bis(3,4,5-trimethoxyphenyl) tetrahydrofuran) and CV-6209 (2-[N-acetyl-N-(2-methoxy-3-octadecyl-carbamoyloxy propoxycarbonyl)aminomethyl]-1-ethylpyridinium chloride) inhibited thapsigargin-induced production of tumor necrosis factor-alpha. The cyclooxygenase inhibitor indomethacin inhibited prostaglandin E2 production, and further enhanced thapsigargin-induced tumor necrosis factor-alpha production in parallel with further increase in cell-associated PAF production. The enhancement of tumor necrosis factor-alpha production induced by thapsigargin plus indomethacin was also inhibited by E6123, L-652,731 and CV-6209. However, exogenously added PAF up to 100 nM did not stimulate production of tumor necrosis factor-alpha. The level of tumor necrosis factor-alpha mRNA was increased by thapsigargin, but was lowered by the PAF receptor antagonist E6123, suggesting that the inhibition of tumor necrosis factor-alpha production by the PAF receptor antagonist is induced at the level of mRNA for tumor necrosis factor-alpha. These findings suggested that concurrently produced cell-associated PAF in thapsigargin-stimulated macrophages up-regulates production of tumor necrosis factor-alpha by acting as an intracellular signaling molecule and the PAF receptor antagonists might penetrate into the cells and antagonize the action of intracellular PAF.

Modulation of early but not later stages of programmed cell death in embryonic avian spinal cord by sonic hedgehog.

Sonic hedgehog (Shh) is a secreted glycoprotein expressed by the notochord and floor plate that is involved in the induction and specification of ventral phenotypes in the vertebrate neural tube. Recently, Shh has also been shown to promote the survival of cultured rat embryo ventral brain and spinal cord cells. We have examined whether Shh can promote the survival of chick embryo neurons in vivo or in vitro. In the chick, Shh is expressed in notochord, floor plate, and ventral neural tube/spinal cord at several stages at which programmed cell death (PCD) occurs. However, the administration of exogenous Shh to embryos in vivo or to motoneuron cultures at these stages failed to promote the survival of several different neuronal populations, including spinal motoneurons, spinal interneurons, sympathetic preganglionic neurons, sensory neurons, and neuronal precursor cells. Rather, at the earliest stage of PCD examined here (embryonic day 3) Shh selectively induced the death of ventral neuronal precursors and floor-plate cells, resulting in a net loss of cells in the neural tube. Altered concentrations of Shh induce aberrant phenotypes that are removed by PCD. Accordingly, normal PCD in the early neural tube may play a role in dorsal-ventral patterning.

A mouse carrying genetic defect in the choice between T and B lymphocytes.

Transgenic mice with human CD3epsilon gene have been shown to exhibit early arrest of T cell development in the thymus. The present study shows that, instead of T cells, B cells are generated in the thymus of a line, tg epsilon26, of the human CD3epsilon transgenic mice. The accumulation of mature B cells in the thymus was found only in tg epsilon26 mice, not in other human CD3epsilon transgenic mouse lines or other T cell-deficient mice, including CD3-epsilon knockout mice and TCR-beta/TCR-delta double knockout mice. Hanging drop-mediated transfer into 2-deoxyguanosine-treated thymus lobes showed that lymphoid progenitor cells rather than thymus stromal cells were responsible for abnormal B cell development in tg epsilon26 thymus, and that tg epsilon26 fetal liver cells were destined to become B cells in normal thymus even in the presence of normal progenitor cells undergoing T cell development. These results indicate that lymphoid progenitor cells in tg epsilon26 mice are genetically defective in thymic choice between T cells and B cells, generating B cells even in normal thymus environment. Interestingly, tg epsilon26 thymocytes expressed GATA-3 and TCF-1, but not LEF-1 and PEBP-2alpha, among T cell-specific transcription factors that are involved in early T cell development, indicating that GATA-3 and TCF-1 expressed during thymocyte development do not necessarily determine the cell fate into T cell lineage. Thus, tg epsilon26 mice provide a novel mouse model in that lineage choice between T and B lymphocytes is genetically defective.

A novel strategy for introducing exogenous bcl-2 into neuronal cells: the Cre/loxP system-mediated activation of bcl-2 for preventing programmed cell death using recombinant adenoviruses.

We have established a novel strategy for introducing exogenous Bcl-2 into neuronal cells that is mediated by Cre/loxP recombination using recombinant adenoviral vectors. An on/off-switching cassette for Bcl-2 (CALNLbcl-2) was designed to express Bcl-2 by recombinase Cre-mediated excisional deletion of a spacer DNA flanked by a pair of loxP sites. Exogenous Bcl-2 was clearly induced in PC12 cell lines carrying CALNLbcl-2 after infection with recombinant adenovirus producing recombinase Cre (AxCANCre). Dual infection with both AxCANCre and a recombinant adenovirus bearing CALNLbcl-2 showed efficient delivery of exogenous Bcl-2 into a hybrid motoneuronal cell line and primary chicken spinal motoneurons. The delivery of foreign Bcl-2 promoted survival of motoneurons in medium either containing or lacking trophic support. Thus, this strategy for delivery of exogenous Bcl-2 will be useful for studying neuronal death as well as for introducing foreign genes into postmitotic neurons under the control of recombinase Cre.

Inhibition of tumor necrosis factor-alpha production by SK&F 98625, a CoA-independent transacylase inhibitor, in cultured rat peritoneal macrophages.

When rat peritoneal macrophages were incubated in medium containing thapsigargin, tumor necrosis factor-alpha (TNF-alpha) production was increased time-dependently. In the presence of SK&F 98625, a CoA-independent transacylase inhibitor, the thapsigargin-induced TNF-alpha production was inhibited dose-dependently. Platelet-activating factor (PAF) and prostaglandin E2 (PGE2) production were also inhibited by SK&F 98625. The SK&F 98625-induced inhibition of TNF-alpha production was not prevented by addition of PGE2. PAF antagonists such as E6123, L-652,731 and CV-6209 partially inhibited the thapsigargin-induced TNF-alpha production, suggesting that concurrently produced PAF in thapsigargin-stimulated macrophages up-regulates TNF-alpha production. The inhibition by SK&F 98625 of thapsigargin-induced TNF-alpha production might be partly due to the inhibition of PAF production.

Mouse Zic1 is involved in cerebellar development.

Zic genes encode zinc finger proteins, the expression of which is highly restricted to cerebellar granule cells and their precursors. These genes are homologs of the Drosophila pair-rule gene odd-paired. To clarify the role of the Zic1 gene, we have generated mice deficient in Zic1. Homozygous mice showed remarkable ataxia during postnatal development. Nearly all of the mice died within 1 month. Their cerebella were hypoplastic and missing a lobule in the anterior lobe. A bromodeoxyuridine labeling study indicated a reduction both in the proliferating cell fraction in the external germinal layer (EGL), from 14 d postcoitum, and in forward movement of the EGL. These findings suggest that Zic1 may determine the cerebellar folial pattern principally via regulation of cell proliferation in the EGL.

A novel type of programmed neuronal death in the cervical spinal cord of the chick embryo.

We examined the massive early cell death that occurs in the ventral horn of the cervical spinal cord of the chick embryo between embryonic days 4 and 5 (E4 and E5). Studies with immunohistochemical, in situ hybridization, and retrograde-tracing methods revealed that many dying cells express Islet proteins and Lim-3 mRNA (motoneuron markers) and send their axons to the somatic region of the embryo before cell death. Together, these data strongly suggest that the dying cells are somatic motoneurons. Cervical motoneurons die by apoptosis and can be rescued by treatment with cycloheximide and actinomycin D. Counts by motoneuron numbers between E3.5 and E10 revealed that, in addition to cell death between E4 and E5, motoneuron death also occur between E6 and E10 in the cervical cord. Studies with [3H]thymidine autoradiography and morphological techniques revealed that in the early cell-death phase (E4-E5), genesis of motoneurons, axonal elongation, and innervation of muscles is still ongoing. However, studies with [3H]thymidine autoradiography also revealed that the cells dying between E4 and E5 become postmitotic before E3.5. Increased size of peripheral targets, treatment with neuromuscular blockade, and treatment with partially purified muscle or brain extracts and defined neurotropic agents, such as NGF, BDNF, neurotrophin-3, CNTF, bFGF, PDGF, S100-beta, activin, cholinergic differentiation factor/leukemia inhibitory factor, bone morphogenetic protein-2, IGF-I, interleukin-6, and TGF-beta 1, were all ineffective in rescuing motoneurons dying between E4 and E5. By contrast, motoneurons that undergo programmed cell death at later stages (E6-E10) in the cervical cord are target-dependent and respond to activity blockade and trophic factors. Experimental approaches revealed that early cell death also occurs in a notochord-induced ectopic supernumerary motoneuron column in the cervical cord. Transplantation of the cervical neural tube to other segmental regions failed to alter the early death of motoneurons, whereas transplantation of other segments to the cervical region failed to induce early motoneuron death. These results suggest that the mechanisms that regulate motoneuron death in the cervical spinal cord between E4 and E5 are independent of interactions with targets. Rather, this novel type of cell death seems to be determined by signals that either are cell-autonomous or are derived from other cells within the cervical neural tube.

Spinovestibular projections in the rat, with particular reference to projections from the central cervical nucleus to the lateral vestibular nucleus.

Projections from the spinal cord to the vestibular nuclei were examined following injections of Phaseolus vulgaris-leucoagglutinin, cholera toxin subunit B, or biotinylated dextran at various levels of the spinal cord in the rat. Labeled terminals were abundant after injections of the tracers into the C2 and C3 segments containing the central cervical nucleus. Labeled terminals were seen in the descending vestibular nucleus and the parvocellular, magnocellular, and caudal parts of the medial vestibular nucleus throughout its rostrocaudal extent. Labeled terminals were most numerous in the lateral vestibular nucleus throughout its rostrocaudal extent. The projections from the central cervical nucleus to the vestibular nuclei were exclusively contralateral to the cells of origin because the axons of the central cervical nucleus neurons cross in the spinal cord. Following tracer injections in the cervical enlargement, many labeled terminals were seen in the magnocellular part of the medial vestibular nucleus, but a few were seen in the lateral and the descending vestibular nucleus. Injections into more caudal segments resulted in sporadic terminal labeling in the magnocellular part of the medial vestibular nucleus, the descending vestibular nucleus, and the caudal part of the lateral vestibular nucleus. The results indicate that primary neck afferent input relayed at the central cervical nucleus is mediated directly to the contralateral vestibular nuclei. It is suggested that this projection serves as an important linkage from the upper cervical segments to the lateral vestibulospinal tract in the tonic neck reflex.

Peptide inhibitors of the ICE protease family arrest programmed cell death of motoneurons in vivo and in vitro.

Members of the CED-3/interleukin-1 beta-converting enzyme (ICE) protease family have been implicated in cell death in both invertebrates and vertebrates. In this report, we show that peptide inhibitors of ICE arrest the programmed cell death of motoneurons in vitro as a result of trophic factor deprivation and in vivo during the period of naturally occurring cell death. In addition, interdigital cells that die during development are also rescued in animals treated with ICE inhibitors. Taken together, these results provide the first evidence that ICE or an ICE-like protease plays a regulatory role not only in vertebrate motoneuron death but also in the developmentally regulated deaths of other cells in vivo.