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Ovarian steroids decrease DNA fragmentation in the serotonin neurons of non-injured rhesus macaques.

Lima FB, Bethea CL - Mol. Psychiatry (2009)

Bottom Line: Two staining patterns were observed, which are referred to as type I, with complete dark staining of the nucleus, and type II, with peripheral staining in the perinuclear area.A montage of the dorsal raphe was created at each level with a Marianas Stereology Microscope and Slidebook 4.2, and the TUNEL-positive cells were counted.In direct comparison with OVX animals, P treatment and E+P treatment significantly reduced the total number of TUNEL-positive cells (Mann-Whitney test, both treatments P=0.04) and E+P treatment reduced the number of TUNEL-positive cells per mm(3) (Mann-Whitney test, P=0.04).

View Article: PubMed Central - PubMed

Affiliation: Division of Reproductive Sciences, Oregon National Primate Research Center, Beaverton, OR 97006, USA.

ABSTRACT
We previously found that ovarian steroids promote neuroprotection in serotonin neurons by decreasing the expression of pro-apoptotic genes and proteins in the dorsal raphe nucleus of rhesus macaques, even in the absence of overt injury. In this study, we questioned whether these actions would lead to a reduction in DNA fragmentation in serotonin neurons. Ovariectomized (OVX) rhesus monkeys were implanted with silastic capsules that were empty (placebo) or containing estradiol (E), progesterone (P) or estradiol and progesterone (E+P) for 1 month. In all animals, eight levels of the dorsal raphe nucleus in a rostral-to-caudal direction were immunostained using the terminal deoxynucleotidyl transferase nick end labeling (TUNEL) method. Two staining patterns were observed, which are referred to as type I, with complete dark staining of the nucleus, and type II, with peripheral staining in the perinuclear area. A montage of the dorsal raphe was created at each level with a Marianas Stereology Microscope and Slidebook 4.2, and the TUNEL-positive cells were counted. In direct comparison with OVX animals, P treatment and E+P treatment significantly reduced the total number of TUNEL-positive cells (Mann-Whitney test, both treatments P=0.04) and E+P treatment reduced the number of TUNEL-positive cells per mm(3) (Mann-Whitney test, P=0.04). Double immunocytochemistry for TUNEL and tryptophan hydroxylase (TPH) indicated that DNA fragmentation was prominent in serotonin neurons. These data suggest that in the absence of ovarian steroids, a cascade of gene and protein expression leads to an increase in DNA fragmentation in serotonin neurons. Conversely, ovarian steroids have a neuroprotective role in the non-injured brain and prevent DNA fragmentation and cell death in serotonin neurons of nonhuman primates.

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Related in: MedlinePlus

Illustration of the anatomical area where the TUNEL staining was counted. Photomicrographs of the montages of 3 levels (out of a total of 8 levels analyzed) of the dorsal raphe nucleus from an ovariectomized monkey treated with placebo that manifest robust TUNEL staining. The montage was built automatically by Slidebook 4.2 and the stereology microscope. Arrows indicate TUNEL positive cells.
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Figure 2: Illustration of the anatomical area where the TUNEL staining was counted. Photomicrographs of the montages of 3 levels (out of a total of 8 levels analyzed) of the dorsal raphe nucleus from an ovariectomized monkey treated with placebo that manifest robust TUNEL staining. The montage was built automatically by Slidebook 4.2 and the stereology microscope. Arrows indicate TUNEL positive cells.

Mentions: In order to perform a stereological analysis of the dorsal raphe nucleus in which positive and negative staining could be segregated, it was necessary to omit the methyl green counterstain. Figure 2 contains montages of representative sections stained by TUNEL at three levels of the dorsal raphe nucleus from an ovariectomized placebo treated animal to illustrate the morphological features.


Ovarian steroids decrease DNA fragmentation in the serotonin neurons of non-injured rhesus macaques.

Lima FB, Bethea CL - Mol. Psychiatry (2009)

Illustration of the anatomical area where the TUNEL staining was counted. Photomicrographs of the montages of 3 levels (out of a total of 8 levels analyzed) of the dorsal raphe nucleus from an ovariectomized monkey treated with placebo that manifest robust TUNEL staining. The montage was built automatically by Slidebook 4.2 and the stereology microscope. Arrows indicate TUNEL positive cells.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2874644&req=5

Figure 2: Illustration of the anatomical area where the TUNEL staining was counted. Photomicrographs of the montages of 3 levels (out of a total of 8 levels analyzed) of the dorsal raphe nucleus from an ovariectomized monkey treated with placebo that manifest robust TUNEL staining. The montage was built automatically by Slidebook 4.2 and the stereology microscope. Arrows indicate TUNEL positive cells.
Mentions: In order to perform a stereological analysis of the dorsal raphe nucleus in which positive and negative staining could be segregated, it was necessary to omit the methyl green counterstain. Figure 2 contains montages of representative sections stained by TUNEL at three levels of the dorsal raphe nucleus from an ovariectomized placebo treated animal to illustrate the morphological features.

Bottom Line: Two staining patterns were observed, which are referred to as type I, with complete dark staining of the nucleus, and type II, with peripheral staining in the perinuclear area.A montage of the dorsal raphe was created at each level with a Marianas Stereology Microscope and Slidebook 4.2, and the TUNEL-positive cells were counted.In direct comparison with OVX animals, P treatment and E+P treatment significantly reduced the total number of TUNEL-positive cells (Mann-Whitney test, both treatments P=0.04) and E+P treatment reduced the number of TUNEL-positive cells per mm(3) (Mann-Whitney test, P=0.04).

View Article: PubMed Central - PubMed

Affiliation: Division of Reproductive Sciences, Oregon National Primate Research Center, Beaverton, OR 97006, USA.

ABSTRACT
We previously found that ovarian steroids promote neuroprotection in serotonin neurons by decreasing the expression of pro-apoptotic genes and proteins in the dorsal raphe nucleus of rhesus macaques, even in the absence of overt injury. In this study, we questioned whether these actions would lead to a reduction in DNA fragmentation in serotonin neurons. Ovariectomized (OVX) rhesus monkeys were implanted with silastic capsules that were empty (placebo) or containing estradiol (E), progesterone (P) or estradiol and progesterone (E+P) for 1 month. In all animals, eight levels of the dorsal raphe nucleus in a rostral-to-caudal direction were immunostained using the terminal deoxynucleotidyl transferase nick end labeling (TUNEL) method. Two staining patterns were observed, which are referred to as type I, with complete dark staining of the nucleus, and type II, with peripheral staining in the perinuclear area. A montage of the dorsal raphe was created at each level with a Marianas Stereology Microscope and Slidebook 4.2, and the TUNEL-positive cells were counted. In direct comparison with OVX animals, P treatment and E+P treatment significantly reduced the total number of TUNEL-positive cells (Mann-Whitney test, both treatments P=0.04) and E+P treatment reduced the number of TUNEL-positive cells per mm(3) (Mann-Whitney test, P=0.04). Double immunocytochemistry for TUNEL and tryptophan hydroxylase (TPH) indicated that DNA fragmentation was prominent in serotonin neurons. These data suggest that in the absence of ovarian steroids, a cascade of gene and protein expression leads to an increase in DNA fragmentation in serotonin neurons. Conversely, ovarian steroids have a neuroprotective role in the non-injured brain and prevent DNA fragmentation and cell death in serotonin neurons of nonhuman primates.

Show MeSH
Related in: MedlinePlus