Calretinin and calbindin-D28K in male rats during postnatal development.

Calcium-binding proteins play potentially important roles in neurogenesis and neuroprotective mechanism(s). Some evidence exists that brain calbindin-D28K (CALB) is regulated by androgens. In the present study, calretinin (CALRET) and CALB patterns were determined by Western analysis in the medial basal hypothalamus (MBH) from male rats along with assaying plasma testosterone levels during postnatal development. Testosterone levels were very low in 7-, 10-, and 30-day-old animals (approximately 0.5 ng/mL), increased in a stair-step fashion to peak levels at 90 days (approximately 3.8 ng/mL), then declined with increasing age to very low levels at 300 days of age (approximately 0.3 ng/mL). At 7 and 10 days, MBH CALRET and CALB levels were low; however, at Day 30 a significant twofold increased was observed. Thereafter, in 60-, 120-, 180-, and 300-day-old animals MBH CALRET and CALB levels were, in general, comparable to 30-day-old values. These findings suggest that there is not a clear correspondence between the androgen status in male rats and the calcium-binding proteins (CALRET & CALB) expressed in the MBH. Therefore, it appears that brain CALRET and CALB are regulated in a developmental fashion with significant increases in expression occurring around the 4th postnatal week.

Aromatase cytochrome P450 and 5 alpha-reductase in the amygdala and cortex of perinatal rats.

The major androgen metabolizing enzymes, aromatase cytochrome P450 and 5 alpha-reductase play critical role(s) in the development of sexually dimorphic brain structures, the modulation of neuroendocrine function(s) and the regulation of sexual and non-sexual behaviors. Using established assays, we detected 5 alpha-reductase and aromatase enzymatic activities in amygdala and frontal cortical tissue from male and female rats during the perinatal interval (from gestational day (GD) 19 to postnatal day (PND) 6). The present findings indicate that 5 alpha-reductase and aromatase rates in the cortex display different enzyme profiles, while in the amygdala tissue site a similar pattern is seen for both enzymes during perinatal development. In general, there was a lack of sex differences in the enzymatic rates. The importance these enzyme systems play in generating androgen (and progesterone) steroid metabolites which influence neural development and function are discussed.

Developmental expression of calretinin in the medial basal hypothalamus and amygdala from male and female rats.

Developmental expression of calretinin in the medial basal hypothalamic (MBH) and amygdala region was examined by Western analysis. Males displayed significantly higher calretinin levels compared to females in the MBH (but not the amygdala) on gestational day 19 and 20. These data imply that hormonal factors may regulate developmental MBH calretinin expression. In turn, sexually dimorphic brain structures might be influenced by calretinin levels that can alter sexually dimorphic patterns of steroidogenesis, cellular migration or programmed cell loss mechanism(s) during neuronal development by modulating intracellular calcium concentrations.

Inhibition of brain 5 alpha-reductase in pregnant rats: effects on enzymatic and behavioral activity.

Medial basal hypothalamic (MBH) 5 alpha-reductase activity was significantly blocked with a known inhibitor, Proscar (Finasteride), in pregnant rats while their open-field behavior was quantified during the last week of pregnancy. In control animals, open-field behavior significantly decreased (in a stair-step fashion) as a function of increasing gestational age. Conversely, in Proscar-treated animals open-field values significantly increased on day 15 and 17 of gestation compared to control values. These data indicate that inhibition of MBH 5 alpha-reductase during pregnancy significantly increased open-field activity levels during late gestation in rats and provides evidence for a link between the production of 5 alpha-reduced metabolites of progesterone in brain and behavioral activity during pregnancy.

Effects of testosterone and progesterone on brain 5alpha-reductase and aromatase in Long-Evans males and comparison of aromatase in Long-Evans vs. Sprague-Dawley rats.

We investigated medial basal hypothalamic-preoptic area (MBH-POA) 5alpha-reductase and aromatase enzyme activities in gonadally intact and castrated adult Long-Evans (L-E) male rats treated with testosterone (T), progesterone (P), and a combination of T+P. MBH-POA 5alpha-reductase and aromatase activities did not differ significantly among the groups. The lack of a difference in MBH-POA aromatase between control and castrated L-E animals was unexpected. In two further experiments, MBH-POA aromatase was examined in intact and castrated L-E and Sprague-Dawley (S-D) rats, using direct and indirect assays. The activity in castrated S-D (but again, not in L-E) rats significantly decreased compared to control values. These data suggest that the absence of gonads does not decrease MBH-POA aromatase in adult L-E rats.

Calbindin-D28k is regulated by adrenal steroids in hypothalamic tissue during prenatal development.

Regulation of calbindin-D28k (CALB) in the medial basal hypothalamus (MBH) from male and female fetuses was examined by Western analysis. Control fetal males displayed significantly higher MBH CALB levels compared to females at gestational day 20. Whereas, in general, the lowest CALB levels were recorded in male and female fetuses from long-term prenatally stressed or fetuses from adrenalectomized pregnant rats. These data indicate that corticosterone regulates MBH CALB expression during prenatal development and CALB may be implicated in modulating the sexual differentiation of neural structures within the MBH during perinatal development.

Hypothalamic aromatase cytochrome P450 and 5alpha-reductase enzyme activities in pregnant and female rats.

The metabolism of steroid hormones in the medial basal hypothalamus (MBH) is known to play a critical role in neural development, the modulation of neuroendocrine function and regulating sexual behavior. While the important biological functions of the aromatase enzyme are well established, the importance of brain 5alpha-reductase has been revealed and elucidated only in the last few years. The distribution and regulation of brain aromatase and 5alpha-reductase enzyme activities have been investigated for the most part in male rats. Therefore, in the present study, MBH aromatase cytochrome P450 and 5alpha-reductase activities were characterized in pregnant and female rats during postnatal development under various hormonal conditions. MBH aromatase activity was determined in each tissue sample using the 'tritiated water release' assay, whereas, the 5alpha-reductase rates were determined by thin layer chromatography and scintillation counting of the isolated 5alpha-metabolites. Both activities were highest in infantile animals, then declined with increasing postnatal age; whereas, in aged non-cycling or ovariectomized/adrenalectomized (Ovx/Adx) rats high rates of androgen metabolism were seen in MBH tissue. No significant alterations in MBH aromatase were observed when the 5alpha-reductase pathway was blocked in pregnant animals during late gestation with a known 5alpha-reductase inhibitor (Proscar). However, plasma estradiol levels were significantly increased in the Proscar-treated animals. These results indicate that: 1) the decreasing MBH aromatase and 5alpha-reductase profile (in infantile to adult cycling animals) is developmentally regulated, 2) evidently, there is a divergent regulatory mechanism controlling MBH aromatase versus 5alpha-reductase in aged animals where the aromatase activity increased in aged non-cycling and Ovx/Adx rats while 5alpha-reductase rates remained at moderate levels and, 3) apparently, the 5alpha-reductase pathway is not involved in regulating MBH aromatase activity during late pregnancy.

Unicompartmental knee replacement after high tibial osteotomy: Invalidating a contraindication.

The outcome of high tibial osteotomy (HTO) deteriorates with time, and additional procedures may be required. The aim of this study was to compare the clinical and radiological outcomes between unicompartmental knee replacement (UKR) and total knee replacement (TKR) after HTO as well as after primary UKR. A total of 63 patients (63 knees) were studied retrospectively and divided into three groups: UKR after HTO (group A; n = 22), TKR after HTO (group B; n = 18) and primary UKR (group C; n = 22). The Oxford knee score (OKS), Knee Society score (KSS), hip-knee-ankle angles, mechanical axis and patellar height were evaluated pre- and post-operatively. At a mean of 64 months (19 to 180) post-operatively the mean OKS was 43.8 (33 to 49), 43.3 (30 to 48) and 42.5 (29 to 48) for groups A, B and C, respectively (p = 0.73). The mean KSS knee score was 88.8 (54 to 100), 88.11 (51 to 100) and 85.3 (45 to 100) for groups A, B and C, respectively (p = 0.65), and the mean KSS function score was 85.0 (50 to 100) in group A, 85.8 (20 to 100) in group B and 79.3 (50 to 100) in group C (p = 0.48). Radiologically the results were comparable for all groups except for patellar height, with a higher incidence of patella infra following a previous HTO (p = 0.02).

Brain aromatase in control versus castrated Norway Brown, Sprague-Dawley and Wistar adult rats.

Brain aromatase cytochrome P450 converts androgens to estrogens that play a critical role in the development of sexually dimorphic neural structures, the modulation of neuroendocrine function(s), and the regulation of sexual behavior. We characterized the influence of surgical castration on brain aromatase in Norway Brown and Wistar adult rats and compared their responses to Sprague-Dawley rats that were surgically or biochemically castrated (with flutamide, a known androgen receptor blocker). Aromata enzyme activity was measured by the tritiated water release assay in the medial basal hypothalmus/preoptic area (MBH/POA) and amygdala brain regions. The present results demonstrate that independent of the rat strain examined, MBH/POA aromatase is regulated by androgens (in Sprague-Dawley, Norway Brown and Wistar males). However, intact Wistar animals displayed significantly higher MBH/POA aromatase levels compared to Sprague-Dawley control values. Conversely, in the amygdala region, there was an apparent lack of androgen hormone action upon aromatase enzyme activity in some of the rat strains tested. The importance of brain aromatase regulating estrogen biosynthesis and influencing brain development and function is covered.

Brain aromatase and 5alpha-reductase, regulatory behaviors and testosterone levels in adult rats on phytoestrogen diets.

The purpose of this study was to examine the short-term effects of phytoestrogens in the diet on regulatory behaviors (food/water intake and locomotor activity), prostate weight, testosterone levels, and brain androgen metabolizing enzyme activity levels in adult male rats. Sprague-Dawley rats were fed phytoestrogen-containing versus phytoestrogen-free diets for 29 days. Standard methods were used to measure open field behavior, reproductive, hormonal parameters, and enzymatic activity levels. The phytoestrogen diet contained approximately 200 microg/g of isoflavones whereas in the phytoestrogen-free diet, no phytoestrogens were detected by HPLC analysis. There were no significant differences in any of the regulatory behaviors (food/water intake or locomotor activity), prostate weight, or testosterone levels between the treatment groups. Furthermore, there was no significant influence of phytoestrogens on brain aromatase activity levels, in either the medial basal hypothalamic-preoptic area (MBH-POA) or amygdala brain tissue sites examined. However, significant alterations in MBH-POA and amygdala 5alpha-reductase activities were detected in animals receiving the phytoestrogen-containing versus the phytoestrogen-free diets.

Neuroendocrine regulation of sexually dimorphic brain structure and associated sexual behavior in male rats is genetically controlled.

Steroid hormones, particularly 17beta-estradiol (E2), regulate the development and expression of neural structures and sexual behavior. Recently, we demonstrated that E2-regulated responses are controlled by quantitative trait loci. In this study, we quantified 1) volume of the sexually dimorphic nucleus (SDN) of the preoptic area (POA); 2) medial basal hypothalamic (MBH)-POA aromatase and 5alpha-reductase enzyme activities during prenatal development and in adults; 3) serum LH, testosterone, FSH, E2, prolactin (PRL), and corticosterone levels; 4) reproductive organ (i.e., testis and ventral prostate) weights; and 5) male mating behavior in Noble (NB/Cr) and Wistar-Furth (WF/NCr) rat strains to determine the genetic influence on the measured parameters. Maximal phenotypic divergence in male SDN-POA volumes was seen between NB/Cr versus WF/NCr and BDIX/Cr rats (among nine rat strains initially examined), with the average SDN-POA volume of NB/Cr male rats being significantly greater ( approximately 30%) than that of either WF/NCr or BDIX/Cr males. Subsequent experiments investigated WF/NCr versus NB/Cr male rats in further detail. Significantly higher MBH-POA aromatase activity was seen in adult WF/NCr versus NB/Cr males, while MBH-POA 5alpha-reductase rates were not significantly different (within or between sex) for the two rat strains assayed. Serum LH levels were significantly higher (by greater than sixfold) in WF/NCr versus NB/Cr males, whereas testis organ:body weight and ventral prostate:body weight ratios in WF/NCr versus NB/Cr males were significantly smaller (by approximately 6-fold for testis and approximately 1.5-fold for prostate values). Serum FSH levels were significantly higher (by twofold) in WF/NCr versus NB/Cr males. However, serum testosterone levels were not significantly different, whereas E2 levels were approximately twofold higher (but not significantly different) in WF/NCr versus NB/Cr animals. No significant differences were found in basal (i.e., nonstress) serum PRL or corticosterone levels between the WF/NCr and NB/Cr males. In male copulatory tests, NB/Cr males exhibited significantly more aggressive sexual behavior (e.g., in mounting, intromission, and ejaculation parameters) compared with WF/NCr males. Taken together, these findings indicate that WF/NCr males are, in general, low responders, whereas NB/Cr males are high responders to hormonal signals. The obtained data suggest that the correlative, phenotypic variation in SDN-POA volume (i.e., structure) and reproductive hormone patterns and mating behavior (i.e., function) of WF/NCr versus NB/Cr males is regulated by potentially E2-mediated mechanisms that are genetically controlled.