TY - JOUR
T1 - Nitric oxide synthase 1 and nitric oxide synthase 3 protein expression is regionally and temporally regulated in fetal brain
AU - Northington, Frances J.
AU - Koehler, Raymond C.
AU - Traystman, Richard J.
AU - Martin, Lee J.
N1 - Funding Information:
We wish to thank Dawn Spicer and Ann Price for expert technical assistance with this study and Karen Boozer for assistance with manuscript preparation. This work was supported in part by USPHS NINDS P01-20020, a Grant in Aid from the American Heart Association, and with funds contributed in part by the AHA, Maryland Affiliate, F.J.N. is supported by a Merck Clinician Scientist Award from Johns Hopkins University School of Medicine and by USPHS NINDS K08-1742-01. L.J.M. is a recipient of a Leadership and Excellence in Alzheimer's Disease (LEAD) award (NIA AG 07914).
PY - 1996/8/20
Y1 - 1996/8/20
N2 - Two constitutively expressed isoforms of nitric oxide synthase (NOS) have been identified, Nos1 and Nos3. Nos1 was originally identified in neuronal cells and Nos3 in endothelial cells. Because the biochemical activity of NOS is developmentally regulated, we tested the hypothesis that protein expression is also developmentally regulated. Antibodies to Nos1 and Nos3 were evaluated for specificity by immunoblotting and then used for immunohistochemistry. In fetal and adult sheep brain homogenates, Nos1 antibodies identified one immunoreactive band of proteins at 155 kDa. The Nos3 antibody detected one immunoreactive band at 145 kDa that comigrated with a reactive band in endothelial cell lysates. Immunoblots of developing neocortex demonstrated that Nos1 was enriched at early gestational ages, whereas Nos3 expression was relatively constant throughout development. By immunohistochemistry, distinct isoform-specific patterns of immunoreactivity were detected. At 60 days, Nos1 immunoreactivity is primarily localized in neuropil, but by midgestation, nonpyramidal neurons are labeled in the cortical plate. Developing neurites are Nos1-positive at 60 and 71 days, decreasing in abundance by 93 days. By 93 days the striatum is fully populated by Nos1-expressing nonprincipal neurons. In hippocampus and subthalamic nucleus, Nos1 immunoreactivity is greatest at 60 and 71 days gestation, decreasing thereafter. Immunoreactivity for Nos3 delineates cerebrovasculature maturation from a primarily radial to a highly complex branching arrangement. Hindbrain structures achieve mature organization of the cerebrovasculature before forebrain. We conclude that constitutive NOS protein expression is developmentally regulated and that distinct isoforms of NOS are regulated differentially during brain development. Expression of Nos3 parallels maturation of the cerebrovasculature, whereas the transient, region- and cell type-dependent enrichment of Nos1 in the developing brain may indicate a temporally and spatially restricted role for this enzyme in the maturation of specific neuronal populations.
AB - Two constitutively expressed isoforms of nitric oxide synthase (NOS) have been identified, Nos1 and Nos3. Nos1 was originally identified in neuronal cells and Nos3 in endothelial cells. Because the biochemical activity of NOS is developmentally regulated, we tested the hypothesis that protein expression is also developmentally regulated. Antibodies to Nos1 and Nos3 were evaluated for specificity by immunoblotting and then used for immunohistochemistry. In fetal and adult sheep brain homogenates, Nos1 antibodies identified one immunoreactive band of proteins at 155 kDa. The Nos3 antibody detected one immunoreactive band at 145 kDa that comigrated with a reactive band in endothelial cell lysates. Immunoblots of developing neocortex demonstrated that Nos1 was enriched at early gestational ages, whereas Nos3 expression was relatively constant throughout development. By immunohistochemistry, distinct isoform-specific patterns of immunoreactivity were detected. At 60 days, Nos1 immunoreactivity is primarily localized in neuropil, but by midgestation, nonpyramidal neurons are labeled in the cortical plate. Developing neurites are Nos1-positive at 60 and 71 days, decreasing in abundance by 93 days. By 93 days the striatum is fully populated by Nos1-expressing nonprincipal neurons. In hippocampus and subthalamic nucleus, Nos1 immunoreactivity is greatest at 60 and 71 days gestation, decreasing thereafter. Immunoreactivity for Nos3 delineates cerebrovasculature maturation from a primarily radial to a highly complex branching arrangement. Hindbrain structures achieve mature organization of the cerebrovasculature before forebrain. We conclude that constitutive NOS protein expression is developmentally regulated and that distinct isoforms of NOS are regulated differentially during brain development. Expression of Nos3 parallels maturation of the cerebrovasculature, whereas the transient, region- and cell type-dependent enrichment of Nos1 in the developing brain may indicate a temporally and spatially restricted role for this enzyme in the maturation of specific neuronal populations.
KW - Brain development
KW - Cerebrovascular development
KW - Constitutive nitric oxide synthase protein
KW - Immunocytochemistry
KW - Sheep
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U2 - 10.1016/0165-3806(96)00051-X
DO - 10.1016/0165-3806(96)00051-X
M3 - Article
C2 - 8873971
AN - SCOPUS:0030595006
SN - 0165-3806
VL - 95
SP - 1
EP - 14
JO - Developmental Brain Research
JF - Developmental Brain Research
IS - 1
ER -