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Neurolin Ig domain 2 participates in retinal axon guidance and Ig domains 1 and 3 in fasciculation.

Leppert CA, Diekmann H, Paul C, Laessing U, Marx M, Bastmeyer M, Stuermer CA - J. Cell Biol. (1999)

Bottom Line: Repeated intraocular injections of a mAb against Ig domain 2 disturb the disk-directed growth: axons grow in aberrant routes and fail to reach the optic disk, but remain fasciculated. mAbs against Ig domains 1 and 3 disturb the formation of tight fascicles. mAb against Ig domain 2 significantly increases the incidence of growth cone departure from the disk-oriented fascicle track, while mAbs against Ig domains 1 and 3 do not.This was demonstrated by time-lapse videorecording of labeled growth cones.Thus, Ig domain 2 of neurolin is apparently essential for growth cone guidance towards the disk, presumably by being part of a receptor (or complex) for an axon guidance component.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Konstanz, D-78457 Konstanz, Germany.

ABSTRACT
The optic disk-directed growth of retinal ganglion cell axons is markedly disturbed in the presence of polyclonal antineurolin antibodies, which mildly affect fasciculation (Ott, H., M. Bastmeyer, and C.A.O. Stuermer, 1998. J. Neurosci. 18:3363-3372). New monoclonal antibodies (mAbs) against goldfish neurolin, an immunoglobulin (Ig) superfamily cell adhesion/recognition molecule with five Ig domains, were generated to assign function (guidance versus fasciculation) to specific Ig domains. By their ability or failure to recognize Chinese hamster ovary cells expressing recombinant neurolin with deletions of defined Ig domains, mAbs were identified as being directed against Ig domains 1, 2, or 3, respectively. Repeated intraocular injections of a mAb against Ig domain 2 disturb the disk-directed growth: axons grow in aberrant routes and fail to reach the optic disk, but remain fasciculated. mAbs against Ig domains 1 and 3 disturb the formation of tight fascicles. mAb against Ig domain 2 significantly increases the incidence of growth cone departure from the disk-oriented fascicle track, while mAbs against Ig domains 1 and 3 do not. This was demonstrated by time-lapse videorecording of labeled growth cones. Thus, Ig domain 2 of neurolin is apparently essential for growth cone guidance towards the disk, presumably by being part of a receptor (or complex) for an axon guidance component.

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Quantification of growth cones departing from their  fascicle in the presence of different antineurolin antibodies. To  quantify the effects caused by the injected neurolin antibodies,  images of all DiO-labeled growth cones found in one retina were  taken. Growth cones elongating directly towards the optic disk  were categorized as directed growth. Growth cones deviating by  >10° from the disk-directed path were categorized as misdirected  growth. The percentage of misdirected growth was determined in  retinae isolated from eyes injected with either mAb N850, mAb  N518, mAb N100, or neurolin Fabs (from a polyclonal antiserum,  pcl Fabs) and compared to control eyes that received either  buffer injections or no injections. mAb N518 (*) or pcl Fabs (*)  caused significantly more misdirected growth than mAb N850 or  mAb N100 (P < 0.001, mAb N518; P < 0.05, pcl Fabs; χ2 test), in  which off-track growth cones were numerically similar to controls.
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Figure 7: Quantification of growth cones departing from their fascicle in the presence of different antineurolin antibodies. To quantify the effects caused by the injected neurolin antibodies, images of all DiO-labeled growth cones found in one retina were taken. Growth cones elongating directly towards the optic disk were categorized as directed growth. Growth cones deviating by >10° from the disk-directed path were categorized as misdirected growth. The percentage of misdirected growth was determined in retinae isolated from eyes injected with either mAb N850, mAb N518, mAb N100, or neurolin Fabs (from a polyclonal antiserum, pcl Fabs) and compared to control eyes that received either buffer injections or no injections. mAb N518 (*) or pcl Fabs (*) caused significantly more misdirected growth than mAb N850 or mAb N100 (P < 0.001, mAb N518; P < 0.05, pcl Fabs; χ2 test), in which off-track growth cones were numerically similar to controls.

Mentions: In buffer-injected and noninjected control retinae, the majority of growth cones (>80%) grew towards the optic disk in association with other labeled (and unlabeled) axons of the same fascicle (Fig. 7). Between 15.5 and 16.9% of the growth cones deviated from the disk-directed route in both types of control retinae (Fig. 7). This relatively high occurrence of growth outside the fascicle may result from the flattening of the retina during preparation, from suboptimal culture conditions, or from labeling and illuminating the growth cones, but is probably not caused by the earlier intraocular injections of fluid, since the amount of off-track growth was similar in retinae of injected and uninjected eyes. In retinae isolated from mAb N850- and mAb N100-injected eyes, the fraction of growth cones deviating from the fascicle track amounted to 22 and 24.1%, respectively, and thus, was similar to that in controls. After neurolin Fab and mAb N518 injections, the fraction of deviant growth cones increased to 47.5 and 51.9%, respectively. The difference between mAb N518, and mAb N850 and N100 effects is statistically significant (P < 0.001, χ2 test). Thus, mAb 518 and neurolin Fabs cause roughly twice as many growth cones to depart from their fascicle as in controls, and in mAb N850- and mAb N100-injected fish. Moreover, as in vivo, the departure of dorsal RGC growth cones from their fascicle was significantly more frequent (n = 21) than that of ventral axons (n = 3). This was determined from 24 deviant growth cones with known positions.


Neurolin Ig domain 2 participates in retinal axon guidance and Ig domains 1 and 3 in fasciculation.

Leppert CA, Diekmann H, Paul C, Laessing U, Marx M, Bastmeyer M, Stuermer CA - J. Cell Biol. (1999)

Quantification of growth cones departing from their  fascicle in the presence of different antineurolin antibodies. To  quantify the effects caused by the injected neurolin antibodies,  images of all DiO-labeled growth cones found in one retina were  taken. Growth cones elongating directly towards the optic disk  were categorized as directed growth. Growth cones deviating by  >10° from the disk-directed path were categorized as misdirected  growth. The percentage of misdirected growth was determined in  retinae isolated from eyes injected with either mAb N850, mAb  N518, mAb N100, or neurolin Fabs (from a polyclonal antiserum,  pcl Fabs) and compared to control eyes that received either  buffer injections or no injections. mAb N518 (*) or pcl Fabs (*)  caused significantly more misdirected growth than mAb N850 or  mAb N100 (P < 0.001, mAb N518; P < 0.05, pcl Fabs; χ2 test), in  which off-track growth cones were numerically similar to controls.
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Related In: Results  -  Collection

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Figure 7: Quantification of growth cones departing from their fascicle in the presence of different antineurolin antibodies. To quantify the effects caused by the injected neurolin antibodies, images of all DiO-labeled growth cones found in one retina were taken. Growth cones elongating directly towards the optic disk were categorized as directed growth. Growth cones deviating by >10° from the disk-directed path were categorized as misdirected growth. The percentage of misdirected growth was determined in retinae isolated from eyes injected with either mAb N850, mAb N518, mAb N100, or neurolin Fabs (from a polyclonal antiserum, pcl Fabs) and compared to control eyes that received either buffer injections or no injections. mAb N518 (*) or pcl Fabs (*) caused significantly more misdirected growth than mAb N850 or mAb N100 (P < 0.001, mAb N518; P < 0.05, pcl Fabs; χ2 test), in which off-track growth cones were numerically similar to controls.
Mentions: In buffer-injected and noninjected control retinae, the majority of growth cones (>80%) grew towards the optic disk in association with other labeled (and unlabeled) axons of the same fascicle (Fig. 7). Between 15.5 and 16.9% of the growth cones deviated from the disk-directed route in both types of control retinae (Fig. 7). This relatively high occurrence of growth outside the fascicle may result from the flattening of the retina during preparation, from suboptimal culture conditions, or from labeling and illuminating the growth cones, but is probably not caused by the earlier intraocular injections of fluid, since the amount of off-track growth was similar in retinae of injected and uninjected eyes. In retinae isolated from mAb N850- and mAb N100-injected eyes, the fraction of growth cones deviating from the fascicle track amounted to 22 and 24.1%, respectively, and thus, was similar to that in controls. After neurolin Fab and mAb N518 injections, the fraction of deviant growth cones increased to 47.5 and 51.9%, respectively. The difference between mAb N518, and mAb N850 and N100 effects is statistically significant (P < 0.001, χ2 test). Thus, mAb 518 and neurolin Fabs cause roughly twice as many growth cones to depart from their fascicle as in controls, and in mAb N850- and mAb N100-injected fish. Moreover, as in vivo, the departure of dorsal RGC growth cones from their fascicle was significantly more frequent (n = 21) than that of ventral axons (n = 3). This was determined from 24 deviant growth cones with known positions.

Bottom Line: Repeated intraocular injections of a mAb against Ig domain 2 disturb the disk-directed growth: axons grow in aberrant routes and fail to reach the optic disk, but remain fasciculated. mAbs against Ig domains 1 and 3 disturb the formation of tight fascicles. mAb against Ig domain 2 significantly increases the incidence of growth cone departure from the disk-oriented fascicle track, while mAbs against Ig domains 1 and 3 do not.This was demonstrated by time-lapse videorecording of labeled growth cones.Thus, Ig domain 2 of neurolin is apparently essential for growth cone guidance towards the disk, presumably by being part of a receptor (or complex) for an axon guidance component.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Konstanz, D-78457 Konstanz, Germany.

ABSTRACT
The optic disk-directed growth of retinal ganglion cell axons is markedly disturbed in the presence of polyclonal antineurolin antibodies, which mildly affect fasciculation (Ott, H., M. Bastmeyer, and C.A.O. Stuermer, 1998. J. Neurosci. 18:3363-3372). New monoclonal antibodies (mAbs) against goldfish neurolin, an immunoglobulin (Ig) superfamily cell adhesion/recognition molecule with five Ig domains, were generated to assign function (guidance versus fasciculation) to specific Ig domains. By their ability or failure to recognize Chinese hamster ovary cells expressing recombinant neurolin with deletions of defined Ig domains, mAbs were identified as being directed against Ig domains 1, 2, or 3, respectively. Repeated intraocular injections of a mAb against Ig domain 2 disturb the disk-directed growth: axons grow in aberrant routes and fail to reach the optic disk, but remain fasciculated. mAbs against Ig domains 1 and 3 disturb the formation of tight fascicles. mAb against Ig domain 2 significantly increases the incidence of growth cone departure from the disk-oriented fascicle track, while mAbs against Ig domains 1 and 3 do not. This was demonstrated by time-lapse videorecording of labeled growth cones. Thus, Ig domain 2 of neurolin is apparently essential for growth cone guidance towards the disk, presumably by being part of a receptor (or complex) for an axon guidance component.

Show MeSH
Related in: MedlinePlus