TY - JOUR
T1 - Ovarian steroid regulation of serotonin-1A autoreceptor messenger RNA expression in the dorsal raphe of rhesus macaques
AU - Pecins-Thompson, M.
AU - Bethea, C. L.
N1 - Funding Information:
Acknowledgements--We thank Dr David Hess, Director of the P30 Hormone Assay Core Laboratory, for the estrogen and progesterone assays, and we are indebted to Dr Eliot Spindel, Director of the P30 Molecular Biology Core Laboratory, for assistance with cloning of the transmembrane domain of the monkey 5-HT1A receptor. We also thank Jared Cooper of the P30 Morphology Core for assistance with cryosectioning. This work was supported by NIH grants HD17269 to C.L.B., DK9098 to M.P.T., P30 Population Center Grant HD18185, and RR00163 for the operation of the ORPRC.
Funding Information:
This study was approved by the Oregon Regional Primate Research Center (ORPRC) Animal Care and Use Committee. Female rhesus monkeys (Maeaca mulatta) were ovariectomized and hysterectomized (spayed) according to previously described procedures 53 three to six months before assignment to this project. Animals ranged in age from six to 15 years old.
PY - 1999/3
Y1 - 1999/3
N2 - It is widely hypothesized that ovarian steroids act on serotonin neurons to modulate mood and alter neuroendocrine function in women. However, information is needed on the molecular consequences of estrogen and progesterone action in serotonin neurons. This study examined the effect of estrogen, with and without progesterone, on the expression of messenger RNA for the serotonin-1A autoreceptor in monkeys using in situ hybridization and a 432-bp serotonin-1A probe generated with polymerase chain reaction. Monkeys were spayed/ovariectomized (control; n=4), estrogen treated (28 days, n=4) and estrogen+progesterone treated (14 days estrogen+14 days estrogen+progesterone, n=4). Perfusion-fixed midbrain sections containing the dorsal raphe (10 μm) were hybridized at 60°C with 35S antisense complementary RNA. After a final wash in 0.1 x standard saline citrate at 70°C, sections were apposed to βmax film for four days and then emulsion fixed. Adjacent sections were immunostained for serotonin to confirm the location of the dorsal raphe. Densitometric analysis of autoradiographs with gray level thresholding was performed at five levels of the dorsal raphe. The number of pixels exceeding background in defined areas was obtained (pixel number), as well as the mean optical density. In the estrogen- and the estrogen+progesterone-treated groups compared to the control group, there was a 38% and 43% decrease in serotonin-1A messenger RNA signal, respectively, represented by pixel number (P<0.05). Mean optical density for serotonin-1A was significantly decreased by estrogen treatment (21%; P<0.05) and then further decreased with the addition of progesterone treatment (45%; P<0.01). Also, the number of positive cells and the grains/cell were counted. There were significantly fewer serotonin-1A messenger RNA-positive cells in the serotonergic neurons of the dorsal raphe in estrogen- and estrogen+progesterone-treated groups (P<0.001) than controls. There were significantly lower single-cell levels of serotonin-1A messenger RNA in serotonergic neurons of the dorsal raphe only in the estrogen+progesterone- treated group (P<0.05). These results suggest that estrogen reduces serotonin-1A gene expression and that the addition of progesterone further reduces serotonin-1A gene expression in non-human primates. If the changes in gene expression are manifested by alterations in protein expression, then, together, these actions of estrogen and progesterone could increase serotonin neurotransmission, thereby elevating mood and/or altering neuroendocrine functions.
AB - It is widely hypothesized that ovarian steroids act on serotonin neurons to modulate mood and alter neuroendocrine function in women. However, information is needed on the molecular consequences of estrogen and progesterone action in serotonin neurons. This study examined the effect of estrogen, with and without progesterone, on the expression of messenger RNA for the serotonin-1A autoreceptor in monkeys using in situ hybridization and a 432-bp serotonin-1A probe generated with polymerase chain reaction. Monkeys were spayed/ovariectomized (control; n=4), estrogen treated (28 days, n=4) and estrogen+progesterone treated (14 days estrogen+14 days estrogen+progesterone, n=4). Perfusion-fixed midbrain sections containing the dorsal raphe (10 μm) were hybridized at 60°C with 35S antisense complementary RNA. After a final wash in 0.1 x standard saline citrate at 70°C, sections were apposed to βmax film for four days and then emulsion fixed. Adjacent sections were immunostained for serotonin to confirm the location of the dorsal raphe. Densitometric analysis of autoradiographs with gray level thresholding was performed at five levels of the dorsal raphe. The number of pixels exceeding background in defined areas was obtained (pixel number), as well as the mean optical density. In the estrogen- and the estrogen+progesterone-treated groups compared to the control group, there was a 38% and 43% decrease in serotonin-1A messenger RNA signal, respectively, represented by pixel number (P<0.05). Mean optical density for serotonin-1A was significantly decreased by estrogen treatment (21%; P<0.05) and then further decreased with the addition of progesterone treatment (45%; P<0.01). Also, the number of positive cells and the grains/cell were counted. There were significantly fewer serotonin-1A messenger RNA-positive cells in the serotonergic neurons of the dorsal raphe in estrogen- and estrogen+progesterone-treated groups (P<0.001) than controls. There were significantly lower single-cell levels of serotonin-1A messenger RNA in serotonergic neurons of the dorsal raphe only in the estrogen+progesterone- treated group (P<0.05). These results suggest that estrogen reduces serotonin-1A gene expression and that the addition of progesterone further reduces serotonin-1A gene expression in non-human primates. If the changes in gene expression are manifested by alterations in protein expression, then, together, these actions of estrogen and progesterone could increase serotonin neurotransmission, thereby elevating mood and/or altering neuroendocrine functions.
KW - 5-HT(1A) autoreceptor
KW - Dorsal raphe
KW - Estrogen
KW - Primate
KW - Progesterone
KW - Serotonin
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U2 - 10.1016/S0306-4522(98)00326-1
DO - 10.1016/S0306-4522(98)00326-1
M3 - Article
C2 - 10051234
AN - SCOPUS:0033105944
SN - 0306-4522
VL - 89
SP - 267
EP - 277
JO - Neuroscience
JF - Neuroscience
IS - 1
ER -