Escherichia coli productora de toxina Shiga: el desafío de adherirse para sobrevivir / Shiga toxin producing Escherichia coli: the challenge of adherence to survive

Resumen Las cepas de Escherichia coli productoras de toxina Shiga (STEC) son reconocidas como responsables de un alto número de casos de enfermedades de transmisión alimentaria a nivel mundial. Su patogenicidad ha sido vinculada directamente con la actividad de las toxinas (Stx); sin embargo, la habilidad de estas bacterias para colonizar al huésped y otras superficies puede ser esencial para desarrollar su poder patogénico. La gran plasticidad genómica de cepas STEC se infiere de la variabilidad de perfiles de virulencia, con la frecuente emergencia de cepas con nuevos genes, codificados en nuevas islas de patogenicidad vinculadas al metabolismo y la adherencia. La formación de biofilm es un mecanismo espontáneo por el cual las cepas STEC resisten en un ambiente hostil, lo que les permite sobrevivir y, de esa forma, llegar al huésped, a través de los alimentos o de las superficies que están en contacto con ellos. Este mecanismo presenta una alta variabilidad intra e interserotipo y su desarrollo no depende solo de los microorganismos que lo conforman. Factores inherentes al ambiente (pH, temperatura) y la superficie (acero inoxidable, poliestireno) a la que pueden adherirse influyen en la expresión de biofilm. El concepto «una salud¼ implica la interrelación entre los actores de salud pública, animal y ambiental para lograr alimentos inocuos y evitar contaminación cruzada y resistencia a sanitizantes, lo cual pone de manifiesto la necesidad de identificar patógenos emergentes a través de nuevos marcadores moleculares, que detecten cepas STEC portadoras del denominado locus for enterocyte effacement (LEE) o del locus de adherencia y autoagregación (LAA).

Abstract Shiga Toxin-producing Escherichia coli (STEC) is recognized as being responsible for a large number of foodborne illnesses around the world. The pathogenicity of STEC has been related to Stx toxins. However, the ability of STEC to colonize the host and other surfaces can be essential for developing its pathogenicity. Different virulence profiles detected in STEC could cause the emergence of strains carrying new genes codified in new pathogenicity islands linked to metabolism and adherence. Biofilm formation is a spontaneous mechanism whereby STEC strains resist in a hostile environment being able to survive and consequently infect the host through contaminated food and food contact surfaces. Biofilm formation shows intra-and inter-serotype variability, and its formation does not depend only on the microorganisms involved. Other factors related to the environment (such as pH, temperature) and the surface (stainless steel and polystyrene) influence biofilm expression. The «One Health¼ concept implies the interrelation between public, animal, and environmental health actors to ensure food safety, prevent cross-contamination and resistance to sanitizers, highlighting the need to identify emerging pathogens through new molecular markers of rapid detection that involve STEC strains carrying the Locus of Enterocyte Effacement or Locus of Adhesion and Autoaggregation.

[Shiga toxin producing Escherichia coli: the challenge of adherence to survive]. / Escherichia coli productora de toxina Shiga: el desafío de adherirse para sobrevivir.

Shiga Toxin-producing Escherichia coli (STEC) is recognized as being responsible for a large number of foodborne illnesses around the world. The pathogenicity of STEC has been related to Stx toxins. However, the ability of STEC to colonize the host and other surfaces can be essential for developing its pathogenicity. Different virulence profiles detected in STEC could cause the emergence of strains carrying new genes codified in new pathogenicity islands linked to metabolism and adherence. Biofilm formation is a spontaneous mechanism whereby STEC strains resist in a hostile environment being able to survive and consequently infect the host through contaminated food and food contact surfaces. Biofilm formation shows intra-and inter-serotype variability, and its formation does not depend only on the microorganisms involved. Other factors related to the environment (such as pH, temperature) and the surface (stainless steel and polystyrene) influence biofilm expression. The «One Health¼ concept implies the interrelation between public, animal, and environmental health actors to ensure food safety, prevent cross-contamination and resistance to sanitizers, highlighting the need to identify emerging pathogens through new molecular markers of rapid detection that involve STEC strains carrying the Locus of Enterocyte Effacement or Locus of Adhesion and Autoaggregation.

Adult CST-null mice maintain an increased number of oligodendrocytes.

The galactolipids galactocerebroside and sulfatide have been implicated in oligodendrocyte (OL) development and myelin formation. Much of the early evidence for myelin galactolipid function has been derived from antibody and chemical perturbation of OLs in vitro. To determine the role of these lipids in vivo, we previously characterized mice lacking galactocerebroside and sulfatide and observed abundant, unstable myelin and an increased number of OLs. We have also reported that mice incapable of synthesizing sulfatide (CST-null) while maintaining normal levels of galactocerebroside generate relatively stable myelin with unstable paranodes. Additionally, Hirahara et al. (2004; Glia 45:269-277) reported that these CST-null mice also contain an increased number of OLs in the forebrain, medulla, and cerebellum at 7 days of age. Here, we further the findings of Hirahara et al. by demonstrating that the number of OLs in the CST-null mice is also increased in the spinal cord and that this elevated OL population is maintained through, at least, 7 months of age. Moreover, we show that the enhanced OL population is accompanied by increased proliferation and decreased apoptosis of oligodendrocytic-lineage cells. Finally, through ultrastructural analysis, we show that the CST-null OLs exhibit decreased morphological complexity, a feature that may result in decreased OL competition and increased OL survival.

Traumatic brain injury induced cell proliferation in the adult mammalian central nervous system.

Recent studies indicate the existence of progenitor cells and their potential for neurogenesis in the subventricular zone (SVZ) and the hippocampus of the normal adult mammalian brain. However, the proliferative response and the specific cell types generated following traumatic brain injury have not been examined. This cellular response to CNS injury was investigated using the fluid percussion injury (FPI) model, a widely accepted rat model that simulates moderate head injury sustained in humans. Forty-eight hours following moderate FPI, adult rats received intraperitoneal injections of the thymidine analogs, 5-bromodeoxyuridine (BrdU) or tritiated thymidine (3H-thymidine), which are markers for mitotic activity. Injured and control animals receiving BrdU were used to determine the total number of cells induced to proliferate. To determine the cellular identity of these proliferating cells, animals receiving 3H-thymidine were sacrificed and sections through the injured area were immunostained with markers for immature and mature astrocytes, activated microglia, neural precursors and mature neurons. These studies showed that the total number of proliferating cells was significantly increased in the injury group for both the SVZ and the hippocampus. However, the proliferating cells in the SVZ did not express any of the cellular markers used, suggesting that they have not yet begun to differentiate. In contrast, there was a significant increase in the number of immature astrocytes and activated microglia, but not neurons, at this early time point in the hippocampus. Taken together, these experiments demonstrate the compensatory capacity of the adult brain to injury and should lead to a new generation of studies aimed at enhancing the neuronal proliferative response.

Normal CNS myelination in transgenic mice overexpressing MHC class I H-2L(d) in oligodendrocytes.

In demyelinating diseases, such as multiple sclerosis, an upregulation of MHC class I expression is thought to contribute to oligodendrocyte/myelin damage. In order to investigate potential physiological consequences of upregulated MHC class I expression in oligodendrocytes, we generated transgenic mice that overexpress H-2L(d) under the control of the proteolipid protein (PLP) promoter (PLP-L(d) mice). We focused our studies on the MHC class I molecule H-2L(d), because of its unique intracellular transport characteristics. In the CNS of PLP-L(d) mice, H-2L(d) was expressed by oligodendrocytes. Furthermore, H-2L(d) protein was transported to and expressed on the surface of oligodendrocytes. Most importantly, this upregulation of MHC class I expression in the CNS of PLP-L(d) mice did not by itself result in a de- or dysmyelinating phenotype. These transgenic mice are likely to provide a unique and novel tool for the analysis of potential roles of MHC class I-mediated mechanisms in demyelinating pathologies.

Purification of oligodendrocytes and their progenitors using immunomagnetic separation and Percoll gradient centrifugation.

In this unit, two techniques are described for the purification of oligodendrocytes and their progenitors from the developing mammalian central nervous system (CNS). The first method utilizes the technique of immunomagnetic separation to selectively isolate oligodendrocytes and their progenitor cells from the optic nerve of prenatal and early postnatal rats. This technique takes advantage of the surface antigens expressed on these cells. A paramagnetic bead is attached to the cells via an antibody bridge. Target cells that are coupled to magnetic beads can then be separated from a heterogeneous cell population using a magnetic field. The second method for isolating oligodendrocytes uses Percoll gradient centrifugation to separate oligodendrocytes from a heterogeneous cell population by virtue of their cell density and allows the direct isolation of oligodendrocytes from animals aged postnatal day 4 (P-4) to adult. This method is particularly useful for assessing physiological systems present in development that may be lost as a result of growing purified neonatal cells in vitro in the absence of neuronal influence.

Growth factor enhanced retroviral gene transfer to the adult central nervous system.

The use of viral vectors for gene delivery into mammalian cells provides a new approach in the treatment of many human diseases. The first viral vector approved for human clinical trials was murine leukemia virus (MLV), which remains the most commonly used vector in clinical trials to date. However, the application of MLV vectors is limited since MLV requires cells to be actively dividing in order for transduction and therefore gene delivery to occur. This limitation precludes the use of MLV for delivering genes to the adult CNS, where very little cell division is occurring. However, we speculated that this inherent limitation of ML V may be overcome by utilizing the known mitogenic effect of growth factors on cells of the CNS. Specifically, an in vivo application of growth factor to the adult brain, if able to induce cell division, could enhance MLV-based gene transfer to the adult brain. We now show that an exogenous application of basic fibroblast growth factor induces cell division in vivo. Under these conditions, where cells of the adult brain are stimulated to divide, MLV-based gene transfer is significantly enhanced. This novel approach precludes any vector modifications and provides a simple and effective way of delivering genes to cells of the adult brain utilizing MLV-based retroviral vectors.

Signals that initiate myelination in the developing mammalian nervous system.

The myelination of axons by oligodendrocytes in the central nervous system and Schwann cells in the peripheral nervous system is essential for the establishment of saltatory conduction. In the absence or destruction of the myelin sheath, as seen in demyelinating diseases, impulse conduction is impeded resulting in severe sensory and motor deficits. Axon myelination is the culmination of a sequence of events that begins with the differentiation of glial cells and proceeds to the transcription and translation of myelin genes, the elaboration of a myelin sheath, and the recognition and ensheathment of axons. This review examines the regulatory mechanisms for each of these steps and compares and contrasts the role of the axon in initiating myelination in the central and peripheral nervous system.

Rapid purification of glial cells using immunomagnetic separation.

By purifying glial cells from brain tissue containing a heterogeneous cell population, a number of interactions that define glial cell diversification and function within the central nervous system have been determined. The current methods for purifying glial cells, however, can be time consuming and costly. In the following study we have adapted the technique of immunomagnetic separation to separately enrich 0-2A progenitor cells and astrocytes from the rat central nervous system (CNS). In this procedure, tissue from the CNS was enzymatically dissociated and incubated in a primary antibody specific to a surface antigen found on the target cell type (e.g. A2B5 or RAN-2). The target cells were then immunologically coupled to magnetic beads, which were precoated with a secondary antibody specific to the primary, and then separated out from the heterogeneous cell population using a magnetic field. We found that the immunomagnetic separation procedure, which was completed within 2 h, produced a near pure population of glial cells (> 99%). This was confirmed by the absence of unbound cells in the bead-bound fraction. The identification and viability of bead-bound cells were established by culturing these cells and subsequently examining their morphology and antigenic expression. This study shows that glial cell types can be separated out of brain tissue to near purity using immunomagnetic separation. This simple procedure is reliable, inexpensive, and achieves levels of purity and viability comparable with currently available techniques of immunopanning and fluorescence-activated cell sorting, within a fraction of the time.

A new Cys2/His2 zinc finger gene, rKr1, expressed in oligodendrocytes and neurons.

The myelination of nerve fibers is essential for the function of the vertebrate nervous system as a prerequisite for fast saltatory conduction of action potentials. In the central nervous system (CNS), myelin is produced by oligodendrocytes. In order to identify gene regulatory proteins involved in the differentiation of this glial cell type or in the expression of myelin-specific genes, we have constructed a cDNA library from a highly enriched population of rat oligodendrocytes and screened this library for members of the Krüppel family of Cys2/His2 zinc finger proteins. One of the identified clones, named rKr1, encodes a novel protein of 650 amino acids which contains 12 carboxy-terminal zinc finger domains and an amino-terminal acidic domain. On Northern blots, a single rKr1 mRNA of 4.3 kb is detected. This message is present in all adult rat tissues tested, with the highest levels found in the CNS and testis. In situ hybridization on the P15 brain revealed that the transcript is expressed in differentiated oligodendrocytes and in subtypes of neurons. Particularly high message levels are found in motor neurons of the brainstem and the spinal cord. The modular structure of the rKr1 protein, in which a potential DNA binding region (the zinc fingers) is combined with a putative activation domain (the acidic region), suggests a function as sequence-specific transcriptional activator.

Molecular and developmental characterization of novel cDNAs of the myelin-associated/oligodendrocytic basic protein.

Several novel myelin-associated/oligodendrocytic basic protein (MOBP) isoforms were identified in this study by cDNA cloning. They are small, highly basic polypeptides comprising 69, 81, and 99 amino acids (8.2, 9.7, and 11.7 kDa, respectively) and show no significant homology with described proteins or domain structures. All (as yet) identified MOBP isoforms are identical in amino acids 1-68 but differ in the length and polarity of the C-terminal region. One isoform, designated MOBP81, was shown to be expressed abundantly during development. Interestingly, MOBP81 has a significant clustering of positively charged residues at positions 69-81, a feature that also has been observed for myelin basic protein (MBP) and Po. As demonstrated by in situ hybridization, MOBP gene expression occurs during development of the rat optic nerve later than that of MBP and proteolipid protein and coincides exactly with the beginning of myelin compaction. The 2.6 kb MOBP81-A transcript is localized in the processes of oligodendrocytes, whereas the 3.8 kb MOBP81-B transcript is restricted to the perinuclear region. Therefore, MOBP81-A and related mRNAs seem to be transported to the periphery of the oligodendrocytes, as is known for the transcripts of the MBP gene. The late developmental expression of the MOBP gene suggests that the MOBP proteins act at the late steps of myelin formation, possibly in myelin compaction and in the maintenance of the myelin sheath.

A new Cys2/His2 zinc finger gene, rKr2, is expressed in differentiated rat oligodendrocytes and encodes a protein with a functional repressor domain.

The function of the vertebrate nervous system is dependent on the proper myelination of its fiber tracts. Myelin of the CNS is produced by oligodendrocytes. To identify gene regulatory proteins expressed in this particular glial cell type, we isolated cDNAs coding for Cys2/His2 zinc finger proteins from a rat oligodendrocyte cDNA library. One clone, named rKr2 (rKr for rat Krüppel-type protein), encodes a protein with 19 carboxy-terminal zinc finger domains and an amino-terminal Krüppel-associated box domain. This amino-terminal domain of the rKr2 protein behaved as a strong transcriptional repressor module when fused to the DNA binding domain of yeast GAL4 and tested on an appropriate reporter construct. High levels of rKr2 mRNA in adult rat tissues were found only in the CNS and testis; in the CNS, the message was predominantly expressed in differentiated oligodendrocytes. The modular structure of the rKr2 protein (carboxy-terminal DNA binding domain, amino-terminal repressor module) and its expression pattern suggest that it acts as a sequence-specific transcriptional repressor in the myelin-producing glial cells of the CNS.

The chronology of oligodendrocyte differentiation in the rat optic nerve: evidence for a signaling step initiating myelination in the CNS.

In order to determine the signals that initiate axon myelination in the CNS, we have chronicled the differentiation of oligodendrocytes in the rat optic nerve and related this to the time course and spatial gradient seen for optic axon myelination. By using markers specific to the varying stages of oligodendrocyte differentiation we found that oligodendrocyte progenitor cells, present throughout the length of the nerve at postnatal day 2, mature into GC+ oligodendrocytes in a chiasm to eye progression. This gradient along the nerve of oligodendrocyte differentiation continues with oligodendrocytes near the chiasm expressing the genes and encoded proteins to MBP and PLP 3 d before oligodendrocytes near the eye. Although oligodendrocyte differentiation and maturation occurs in a chiasm to eye gradient along the nerve, optic axon segments near the eye are ensheathed with myelin before segments near the chiasm. This suggests that the myelination of optic axons is initiated by a signaling step that is independent of oligodendrocyte differentiation and is stronger near the eye than the chiasm region of the nerve. By examining proposed axonal signals, we found that the onset of myelination is independent of the electrical activity of an axon but can be correlated to the size of an axon.

Developmental expression of the prion protein gene in glial cells.

Replication of prions is dependent on the presence of the host protein PrPc. During the course of disease, PrPc is converted into an abnormal isoform, PrPSc, which accumulates in the brain. Attempts to identify the cell type(s) in which prion replication and PrP conversion occur have reached conflicting results. Although PrP mRNA is present in high amounts in neurons throughout the life of the animal, PrPSc initially accumulates in astrocytes and possibly other glial cells and, later in the course of the disease, spreads diffusely in the tissue, often in white matter. We report here that PrP mRNA is expressed not only in neurons but also in astrocytes and oligodendrocytes throughout the brain of postnatal hamsters and rats. The level of glial Prp mRNA expression in neonatal animals was comparable to that of neurons and increased two-fold during postnatal development. A substantial portion of brain PrP mRNA is therefore contributed by glial cells. Our results provide an explanation for the accumulation of PrPSc in white matter tissue and in the cytoplasm of glial cells and argue for a direct involvement of glia in prion propagation.

A role for oligodendrocytes in the stabilization of optic axon numbers.

Differentiated oligodendrocytes express neurite growth inhibitory proteins at a time when these cells are involved in the myelination of recently formed fiber pathways. As the process of myelination follows the completion of neurite outgrowth and is concurrent with the stabilization of fiber numbers in a pathway, we set out to determine whether myelination and fiber tract stability could be causally related. Myelin formation was prevented in the rat retinofugal pathway by x-irradiating the optic nerves during oligodendrocyte proliferation. Electron microscopic and immunohistochemical analysis of irradiated optic nerves at P15 showed that oligodendrocytes and myelin were virtually absent. Optic fiber numbers were determined at 2 weeks of age throughout the length of normal and x-irradiated nerves. In some cases, normal or irradiation-treated pups were intraocularly injected with FGF 5 d prior to the fiber count in order to promote neurite outgrowth. Axon counts showed that the total fiber number in a myelin-free optic nerve was 10-30% higher than that of a myelinated nerve. Further, fiber numbers fluctuated by as much as 20% along the length of a myelin-free nerve but were relatively constant throughout the length of a myelinated nerve. Treatment of myelinated nerves with fibroblast growth factor (FGF) had no effect on either total fiber numbers or fiber number fluctuation. Conversely, fiber numbers in myelin-free/FGF-treated optic nerves were as much as 40% higher than in normals. Furthermore, total fiber numbers along the length of these nerves fluctuated by up to 34%. These results indicate that, in the absence of myelination, optic fibers are able to form sprouts.(ABSTRACT TRUNCATED AT 250 WORDS)

The role of oligodendrocytes and myelin on axon maturation in the developing rat retinofugal pathway.

In neonatal mammals, newly grown optic axons are uniformly small in diameter. In the adult, in contrast, axons within the optic nerve can be classified into distinct groups according to their diameter. Because axon diameters are also related to the thickness of the myelin sheath, which in turn determines the velocity of action potential propagation, the question of what determines the axon diameter is of critical importance. In a project aimed at determining the influence of the ensheathing cell on axon maturation, oligodendrocyte development was prevented by eliminating their precursors by unilateral x-irradiation at birth. Axon diameters in both the normal and the myelin-free optic nerves were then measured at varying stages of development. The results demonstrate that axon diameter growth remained substantially reduced in the absence of oligodendrocytes. Interestingly, by x-irradiating the optic nerve and tract on one side of the brain, fibers crossing the chiasm became larger as they went from an unmyelinated nerve to a myelinated tract; fibers on the nonirradiated side became smaller as they went from a myelinated nerve and crossed into the nonmyelinated tract. These results clearly point to a local regulation of axon diameter by oligodendrocytes. Moreover, ganglion cells measured 9 d after the initiation of myelination (postnatal day 6, P6) were of similar size within normal retinas and retinas whose axons were x-irradiated, suggesting that ganglion cell growth occurs in spite of the lack of myelin and axon diameter maturation. Finally, we showed, through both section staining with antibodies to myelin basic protein (MBP) and Northern blot analysis using a probe to MBP, that the x-irradiated nerve began a delayed myelination period (in a gradient from chiasm to eye) at P15 and reached an almost normal myelin pattern at P28. Axons from these nerves grew to seemingly normal diameter concomitant with this delayed myelination.

Observations on the early development of the optic nerve and tract of the mouse.

The early development of the retinofugal pathway of mice has been studied by light and electron microscopic methods in order to define the spatial distribution and the structure of the growth cones as they advance from the eye to the brain. We have studied the relationships of the growth cones to each other, to the glia and, in the older individuals, to the nerve fibers that are already terminating in the brain. We have looked at the rate of advance of the growth cones and have paid particular attention to the changing relationships of the growth cones as they approach the optic chiasm. We have also looked to see whether, at early stages, it is possible to recognise any characteristic features distinguishing the fibers destined to be the thickest in the adult, which come from ganglion cells that are generated among the earliest ganglion cells. In transverse sections through the optic stalk about 50-100 microns behind the eye, the first bundles of fibers are seen on embryonic day 12.5 (E12.5) as a mixture of thin (less than 0.5 micron) axons, thicker growth cones, and fine filopodial and foliopodial extensions. During the next two days, as these bundles in the intraorbital nerve increase in size and number, growth cones can be seen in all of the bundles and in all parts of the bundles. They show only a slight preference for one part of the nerve relative to another, and our material provides no evidence for the view that axons are particularly inclined to follow pre-existing bundles. The structure of the pathway changes significantly as it is traced towards the chiasm, and no section or small stretch of sections can be regarded as representative of the nerve as a whole. As the fibers approach the optic chiasm the growth cones come to lie predominantly close to the pial surface, with the deeper regions occupied almost entirely by fine axons. The change occurs in a region where the glial environment also changes, and where a characteristic neural tube-like organization first becomes recognizable. Here the glial cells lie in a periventricular position and send slender radial processes out towards the subpial surface. The newly invading axons in the early optic nerve taper from a broad growth cone back to an extremely slender axon, less than 0.5 micron in diameter. The tapered region is of the order of 100-300 microns in length and advances through the nerve at approximately 60 microns per hour.(ABSTRACT TRUNCATED AT 400 WORDS)

Neurogenesis in the retinal ganglion cell layer of the rat.

The present study has examined the birthdates of neurons in the retinal ganglion cell layer of the adult rat. Rat fetuses were exposed to tritiated thymidine in utero to label neurons departing the mitotic cycle at different gestational stages from embryonic days 12 through to 22. Upon reaching adulthood, rats were either given unilateral injections of horseradish peroxidase into target visual nuclei in order to discriminate (1) ganglion cells from displaced amacrine cells, (2) decussating from non-decussating ganglion cells, and (3) alpha cells from other ganglion cell types; or, their retinae were immunohistochemically processed to reveal the choline acetyltransferase-immunoreactive amacrine cells in the ganglion cell layer. Retinae were embedded flat in resin and cut en face to enable reconstruction of the distribution of labelled cells. Retinal sections were autoradiographically processed and then examined for neurons that were both tritium-positive and either horseradish peroxidase-positive or choline acetyltransferase-positive. Tritium-positive neurons in the ganglion cell layer were present in rats that had been exposed to tritiated thymidine on embryonic days E14-E22. Retinal ganglion cells were generated between E14 and E20, the ipsilaterally projecting ganglion cells ceasing their neurogenesis a full day before the contralaterally projecting ganglion cells. Alpha cells were generated from the very outset of retinal ganglion cell genesis, at E14, but completed their neurogenesis before the other cell types, by E17. Tritium-positive, horseradish peroxidase-negative neurons in the ganglion cell layer were present from E14 through to E22, and are interpreted as displaced amacrine cells. Choline acetyltransferase-positive displaced amacrine cells were generated between E16 and E20. Individual cell types showed a rough centroperipheral neurogenetic gradient, with the dorsal half of the retina slightly preceding the ventral half. These results demonstrate, first, that retinal ganglion cell genesis and displaced amacrine cell genesis overlap substantially in time. They do not occur sequentially, as has been commonly assumed. Second, they demonstrate that the alpha cell population of retinal ganglion cells and the choline acetyltransferase-immunoreactive population of displaced amacrine cells are each generated over a limited time during the periods of overall ganglion cell and displaced amacrine cell genesis, respectively. Third, they show that the very earliest ganglion cells to be generated in the temporal retina have exclusively uncrossed optic axons, while the later cells to be generated therein have an increasing propensity to navigate a crossed chiasmatic course.

Cell Survival in the Uncrossed Projection of the Mammalian Retina is Independent of Birthdate.

Naturally occurring cell loss in the retinal ganglion cell population of one eye can be interrupted by removal of the other eye in newborn rodents. Many of the rescued retinal ganglion cells which project ipsilaterally reside in the nasal retina, that part of the retina normally giving rise to primarily crossed optic axons. Their naturally occurring elimination has been attributed to their hypothesized late neurogenesis and the consequent delayed time of arrival of their axons in the target visual nuclei, thereby placing them at a competitive disadvantage with other, early arriving, optic axons. By combining the technique of tritiated thymidine autoradiography with the retrograde axonal transport of horseradish peroxidase in rats that had been enucleated on the day of birth, we report here that rescued cells in the nasal retina which project ipsilaterally are generated at the same time as their neighbours in the temporal retina. Time of genesis does not distinguish them; consequently, their axons should not differ in their arrival times within the target visual nuclei. Since their only obvious anomaly is one of pathway choice at the optic chiasm, their place of arrival, rather than their time, may ultimately determine their naturally occurring elimination.