HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by TORRES, J. M. Articles by ORTEGA, E. Search for Related Content PubMed PubMed Citation Articles by TORRES, J. M. Articles by ORTEGA, E.

Differential regulation of steroid 5-reductase isozymes expression by androgens in the adult rat brain

J. M. TORRES^* and E. ORTEGA^*,,1

^* Department of Biochemistry and Molecular Biology, Faculty of Medicine, and
Institute of Neurosciences, University of Granada, 18012 Granada, Spain

¹Correspondence: Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Granada, Avda. de Madrid s/n, 18012 Granada, Spain. E-mail: esortega{at}ugr.es

	ABSTRACT

TOP ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
The enzyme 5-reductase (5-R) is present in many mammalian tissues, including the brain. The physiological importance of 5-R in the brain derives from its capability to convert testosterone (T) to a more potent androgen, dihydrotestosterone (DHT), and to convert progesterone and deoxycorticosterone (DOC) to their respective 5-reduced derivatives, precursors of allopregnanolone and tetrahydroDOC, potent allosteric modulators of the -aminobutyric acid receptor (GABA_A-R). 5-R occurs as two isoforms, 5-R type 1 (5-R1) and 5-R type 2 (5-R2). We studied the effects of T and DHT on the mRNA levels of both 5-R isozymes in the prefrontal cortex of the adult rat, using an accurate and precise method that combines the high specificity of one-step quantitative RT-PCR with the sensitivity of capillary electrophoresis. Our results demonstrate that both isozymes of 5-R are expressed in the cerebral cortex of adult rats. The gene expression of 5-R type 2 is under the positive control of T and DHT. The gene that codes for 5-R type 1 is not constitutive, because its expression is negatively regulated by T and DHT. These results open up a new research line that may lead to a better understanding of the role of 5-R isozymes in the physiology of the central nervous system.—Torres, J. M., Ortega, E. Differential regulation of steroid 5-reductase isozymes expression by androgens in the adult rat brain.

Key Words: transcriptional regulation • prefrontal cortex

	INTRODUCTION

TOP ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
THE ENZYME 5-REDUCTASE (5-R) (EC 1.3.99.5) is present in many mammalian tissues, including the brain. The physiological importance of 5-R in the brain may derive from two of its properties: its capability to convert testosterone (T) to a more potent androgen, dihydrotestosterone (DHT), which appears to participate in the sexual differentiation processes of some brain regions (1 2 3 4) ; and its capability to convert progesterone and deoxycorticosterone (DOC) to their respective 5-reduced derivatives, providing the substrates for 3-hydroxysteroid dehydrogenase, the enzyme responsible for the formation of 3-hydroxy-5-pregnan-20-one (allopregnanolone), and 3,21-dihydroxy-5-pregnan-20-one (tetrahydroDOC). These compounds are potent neurosteroids whose action is mediated by allosteric modulation of the -aminobutyric acid–A receptor complex (GABA_A-R) (5) . Indeed, neurosteroids also regulate the expression of genes that encode subunits of GABA_A-R (6) . GABA_A-R participates in the regulation of various psychophysiological phenomena. Thus, fluctuations in the concentrations of these 5-reduced neurosteroids may play a major role in the physiology of the individual and in the development of some disorders (7 8 9 10 11 12 13 14 15 16 17 18) .

5-R exists as two isoforms, 5-R type 1 (5-R1) and 5-R type 2 (5-R2). These two isozymes have different biochemical properties and may therefore be expected to have different physiological roles. In the central nervous system (CNS) of the rat, 5-R type 1 is constantly present at all stages of development and throughout the life of the individual, whereas 5-R type 2 is expressed during the perinatal period and its subsequent expression is confined to specific brain areas (1) . It has been suggested that 5-R type 2 participates in sexual differentiation processes in the CNS, whereas 5-R type 1 essentially plays a catabolic and neuron protective role (1) . 35-reduced neurosteroids regulate physiological processes throughout life. Furthermore, DHT may be necessary to maintain functions previously established during ontogenesis of the CNS. For this reason, both isozymes of 5-R may play an important role in the physiology and pathology of the CNS not only during ontogenesis, but also throughout the individual’s life.

The present paper aimed to study the expression of both isozymes of 5-reductase and their regulation by T and DHT in prefrontal cortex of adult rats, using a novel method developed by our group that combines the high specificity of one-step quantitative RT-PCR with the sensitivity of laser-induced fluorescence capillary electrophoresis (LIF-CE).

	MATERIALS AND METHODS

TOP ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Animals
Adult male Wistar rats weighing 260–280 g were housed in an air-conditioned room with fluorescent lights on from 7.00 to 19.00 h and given standard laboratory pellet chow and water ad lib. Experiments were made in strict accordance with the NIH guide for the "Care and Use of Laboratory Animals." The experimental groups studied were intact rats (I), intact rats plus T (I+T), intact rats plus DHT (I+DHT), castrated rats (C), castrated rats plus T (C+T), and castrated rats plus DHT (C+DHT). The castrated animals underwent bilateral orchidectomy under ether anesthesia. Groups I+T and C+T were subcutaneously (s.c.) injected with oil vehicle (20% ethanol in sesame oil) containing testosterone propionate (T_p; 1 mg/kg body weight/day) (19) on days 0, 3, 6, 9, and 12; a final injection was given 3 h before decapitation on day 15. To enable comparison of the effects of T and DHT, groups I+DHT and C+DHT were s.c. injected with oil vehicle (20% ethanol in sesame oil) containing dihydrotestosterone propionate (D_p; 1 mg/kg body weight/ day) (19) on the same days (days 0, 3, 6, 9, 12, and 15). I and C groups were s.c. injected on the same days with oil vehicle alone. The number of rats per group was 10. The animals were decapitated and the brain was removed and weighed. Prefrontal cortex samples were frozen in liquid nitrogen and stored at -80°C until analysis. Blood samples were collected in heparinized tubes. After coagulation, the blood was centrifuged at 2000 rpm for 10 min. The plasma was separated and stored at -80°C until hormonal measurements were performed.

Hormone assays
Plasma T concentrations were measured by RIA using a commercial DiaSorin (Vercelli, Italy) kit without modification. The intra- and interassay coefficients of variation were 7.6% and 12.0%, respectively, and the sensitivity was 0.05 ng/mL. Plasma DHT concentrations were measured by direct ELISA (Diagnostic Biochem Canada Inc., Ontario, Canada). The intra- and interassay coefficients of variation were 5.9% and 7.5%, respectively, and the sensitivity was 6.0 pg/mL.

Oligonucleotides used for amplifications
Sequences of rat 5-R isozymes were obtained from GeneBank® and the sequence of plasmid pEGFP-C1 was obtained from the Clontech web page. These sequences were used to design the primer pairs. Primers for 5-R isozymes were 20 bp in length, whereas primers used to synthesize both competitor molecules were 40 bp in length. All forward primers were end-labeled with 6-carboxy-fluorescein (6-FAM). Oligonucleotides were synthesized by PE Applied Biosystems, U.K. Primer sequences (5'-3') and PCR product sizes were as shown in Table 1 .

Construction of the internal standard template
Two synthetic internal standard (IS) DNAs of 300 bp were synthesized from the sequence of plasmid pEGFP-C1 (Clontech, Palo Alto, CA, USA). Two different sets of primers of 40 nucleotides were used to synthesize both competitive molecules: IS-1 (competitor DNA of 5-R1) and IS-2 (competitor DNA of 5-R2). In each case, the first 20 nucleotides at the 5' ends correspond to the forward and reverse primers that amplify 5-R1 and 5-R2 fragments, respectively; 20 nucleotides at the 3' ends correspond to the opposite strands of the plasmid sequence. Both 300 bp fragments IS-1 and IS-2 were obtained after two consecutive amplifications from pEGFP-C1, with 5' and 3' ends modified to contain the same nucleotide sequences as SRD5A1 or SRD5A2, following Torres et al. (20) .

Reverse transcription reaction-polymerase chain reaction
Total RNA was extracted from 25 mg of rat prefrontal cortex tissues by acid-guanidinium thiocyanate-phenol-chloroform (21) . The RNA was resuspended in diethyl pyrocarbonate (DEPC) -treated water and quantitated spectrophotometrically for analysis. First-strand cDNA was synthesized from 100 ng of total RNA using the reverse primer and Tth DNA polymerase (GeneCraft, Germany). Reactions were incubated at 55°C for 10 min, followed by 20 min at 70°C. The percentage efficiency for RT reactions was estimated to be 9 3 ± 1.9 (mean±SE for n=7) and was calculated using the formula % RT= ng cDNA synthesized / ng RNA, making the RT reaction in the presence of 10 µCi [-³²P] dCTP (3000 Ci/mmol; 10 mCi/mL). The PCR profile was denaturing, 94°C for 30 s; annealing, 55°C for 30 s; and extension, 72°C for 30 s. In each case the number of cycles was 35. PCR was carried out in a Perkin-Elmer 2400 Thermal Cycler.

Analysis of PCR products
Capillary electrophoresis was carried out in a 47 cm silica capillary containing POP-4 polymer (PE Applied Biosystems). Each sample was run for 24 min at 60 KJ of voltage, with 5 s of injection time. We performed LIF-CE in an ABIPRISM 310 Genetic Analyzer (PE Applied Biosystems).

Statistical analysis
Statistical analysis of the results was performed using the Student’s t test. The data are expressed as mean ± SE.

	RESULTS

TOP ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Serum hormonal levels
Castrated animals had significantly lower T levels than intact animals (Fig. 1 ). As could be expected, there was a significant increase in T levels after T treatment in intact and castrated rats compared with their pretreatment levels. After DHT treatment, there was a significant decrease in T levels in intact rats compared with their pretreatment levels.

View larger version (38K): [in this window] [in a new window]   Figure 1. Effects of testosterone (T) and dihydrotestosterone (DHT) on plasma T levels of intact (I) and castrated (C) animals. *P < 0.01 or less vs. I animals. #P < 0.01 or less vs. C animals.

The DHT levels in castrated animals were lower than those in intact animals (Fig. 2 ). After T treatment, there was a significant increase in DHT levels in both intact and castrated animals compared with their pretreatment levels, with a higher effect in the castrated rats. After DHT treatment, there was an increase in DHT levels in intact and castrated animals compared with their respective pretreatment levels.

View larger version (43K): [in this window] [in a new window]   Figure 2. Effects of testosterone (T) and dihydrotestosterone (DHT) on plasma DHT levels of intact (I) and castrated (C) animals. *P< 0.01 or less vs. I animals. #P < 0.01 or less vs. C animals.

Quantification of 5-R1 mRNA levels in prefrontal cortex
The amount of mRNA was expressed as number of mRNA copies per 100 ng of total RNA. After cDNA was generated from total RNA by RT reaction, it was coamplified in the presence of decreasing amounts of the competitive DNA (64x10⁶–0.5x10⁶ molecules). We coamplified 5-R1 cDNA and the competitive standard DNA IS-1 using the same pair of primers. With decreasing amounts of the competitive DNA, the relative intensity of amplified product of target DNA increased.

The ratio of fluorescence of 5-R1/IS-1 was plotted against the amount of competitive DNA IS-1 and the concentration of target DNA in the sample was calculated according to Torres et al. (20) . The concentration of problem cDNA was corrected by the correction factor K. The correction factor K depends on the RT-PCR characteristics and is the product of three components that represent the correction due to the difference in size between problem and standard, the correction due to the addition of the internal standard in DNA form, and the efficiency of retrotranscription (20) .

The mean amount of 5-R1 mRNA in the prefrontal cortex of the different experimental groups is displayed in Fig. 3 . 5-R1 mRNA levels in castrated animals were significantly higher than in intact animals. After T and DHT treatment, there was not a significant increase in 5-R1 mRNA levels in intact rats compared with their respective pretreatment levels. After T and DHT treatment, there was a significant decrease in 5-R1 mRNA levels in castrated animals compared with their respective pretreatment levels, with a greater effect in the latter group.

View larger version (42K): [in this window] [in a new window]   Figure 3. Effects of testosterone (T) and dihydrotestosterone (DHT) on steroid 5-reductase type 1 (5-R1) mRNA levels of intact (I) and castrated (C) animals in prefrontal cortex of adult male rat. *P < 0.01 or less vs. I animals. #P < 0.01 or less vs. C animals.

Quantification of 5-R2 mRNA levels in prefrontal cortex
In the same way, we coamplified 5-R2 cDNA and the competitive standard DNA IS-2 using the same pair of primers. With decreasing amounts of the competitive DNA, the relative intensity of amplified product of target DNA increased. Thus, the ratio of fluorescence of 5-R2 /IS-2 was plotted against the amount of competitive DNA IS-2. The mean amount of 5-R2 mRNA in the prefrontal cortex of the different experimental groups is shown in Fig. 4 . Castrated animals had significantly lower 5-R2 mRNA levels than intact animals. After T treatment, there was a significant increase in 5-R2 mRNA levels in both intact and castrated animals compared with their pretreatment levels, with a greater effect in the latter group. After DHT treatment, there was a significant increase in 5-R2 mRNA levels in both intact and castrated animals vs. their respective pretreatment levels.

View larger version (34K): [in this window] [in a new window]   Figure 4. Effects of testosterone (T) and dihydrotestosterone (DHT) on steroid 5-reductase type 2 (5-R2) mRNA levels of intact (I) and castrated (C) animals in prefrontal cortex of adult male rat. *P <0.01 or less vs. I animals. #P <0.01 or less vs. C animals.

	DISCUSSION

TOP ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
The results of our experiment clearly demonstrate that both isozymes of 5-R are expressed in prefrontal cortex of adult male rats. It was previously reported that 5-R type 2 is selectively expressed in the hypothalamus of adult male rats under basal conditions (22) . By using a novel method of "one-step" quantitative RT-PCR coupled to LIF-CE, we also detected 5-R type 2 expression in the prefrontal cortex of adult rats under basal conditions. It is therefore possible that 5-R type 2 is expressed in many areas of the brain of adult rat but has not yet been detected because of methodological limitations, as was disclosed when our group used a two-step quantitative RT-PCR coupled to CE in an earlier study (20) .

The pattern of expression of 5-R type 2 presented in this study appears to be correlated with T levels, indicating that this isozyme may be positively modulated by androgen, as described previously during late fetal/early post-natal life (1) . Our results demonstrate for the first time that 5-R type 2 expression in the brain is regulated by DHT, a product of the encoded enzyme, via a form of feed-forward regulation. This regulatory mechanism is common among developmentally important genes (19 , 23) . Thus, T and DHT bind to a transcription factor, the androgen receptor, which in turn activates genes that carry out morphogenetic programs implicated in sexual differentiation processes in specific brain regions (1) . In view of our results in adult animals, 5-R type 2 may play a role in the CNS not only during its ontogenesis but also during the lifespan of the individual, as occurs in the prostate (24) .

Our study demonstrated that 5-R type 1 expression is not controlled by androgen in the intact animal. However, it is negatively controlled by some testicular factors (in part by T and DHT) because there is a marked increase in 5-R type 1 expression in castrated animals, in which circulating T and DHT are low. Furthermore, T and DHT decrease the mRNA levels of 5-R type 1 in castrated animals. These findings are novel and surprising. Previous reports have attributed an essentially catabolic role to 5-R type 1 (13) , protecting neurons from excessive glucocorticoids that may induce apoptotic processes (25 , 26) . This is not unreasonable because 5-R type 1 actively metabolizes steroids when they reach high concentrations within cells (27) . From a physiological standpoint, the elevated expression of 5-R type 1 in the castrated animal, when steroid levels are lower than in the intact animal, suggests that the catabolic function is not the only role played by this isozyme. The expression profile of 5-R type 1 in the cerebral cortex of the adult male rat is totally different from its profile in the liver (unpublished data), which is the catabolic organ par excellence.

Our results raise the possibility that 5-R type 1 may also be involved in sexual dimorphism processes, exerting effects opposite to those of 5-R type 2, given that the two isozymes are regulated in opposite directions by T and DHT. 5-R type 1 may also participate in the synthesis of 5-reduced derivatives of progesterone, especially in females in whom progesterone levels are high (28 , 29) or in stress situations. It has been reported that in stress situations the ratio allopregnanolone/progesterone increases in the cerebral cortex of both intact and adrenalectomized male rats (7) , pointing that allopregnanolone biosynthesis occurs in situ in the brain from progesterone and that stress situations increase the via of the 5-reduction in the brain.

We previously reported that GABA_A receptors participate in the development of gender differences in the brain (30) . Thus, the neonatal administration of the GABA agonist diazepam (DZ) to male rats induced maternal behaviors and decreased the number of accessory olfactory bulb (AOB) mitral cells in adulthood to values found in the female (31) . 35-reduced neurosteroids regulate GABA_A receptors in a similar way to barbiturates (17 , 32) and may therefore exert similar effects in the CNS. We found that prenatal environmental stress altered sex differences in parental behavior and increased AOB mitral cells of the rats in adulthood (unpublished data). Stress situation increased allopregnanolone levels in the cerebral cortex of rats (7) and perhaps in other cerebral areas. 5-R activity has been demonstrated in several brain areas, including the olfactory bulb (33) . On the basis of these findings, 35-reduced neurosteroids, and therefore 5-R type 1, may be involved (besides other functions) in the sexual differentiation of the brain and of behavior, favoring the formation and maintenance of female brain structures. Our data demonstrated an increase in 5-R type 1 expression in the castrated male rat that is hormonally similar to a female model. This observation is consistent with a previous report that 5-R activity in liver, which expresses only 5-R type 1, is more active in female than in male rats (34) .

Earlier findings have demonstrated that neonatal administration of DHT induced a decrease in the volume of AOB in male rat to levels found in females (35) . It is difficult to understand how DHT, an essentially androgenic hormone, can feminize brain structures. A reasonable explanation may be that the exogenously administered DHT overexpressed 5-R type 2 in the brain in a generalized manner, producing 35-reduced metabolites that have the same effect as DZ on AOB volume (31) . It has been reported that 5-R type 2 gene expression was induced in the hippocampus of female mice by progesterone producing 5-reduced neurosteroids, like allopregnanolone, which are thought to exert behavioral effects such as anxiolytic and anesthetic (36) . The contribution of 5-R type 2 to the formation of 35-reduced neurosteroids may be small in the male brain under physiological conditions because concentrations of T are much higher than those of progesterone or DOC. Furthermore, the male brain may avoid the generalized overproduction of DHT from T. In fact, T induces the neuronal enzyme aromatase in the brain (22 , 37) , and it is well known that most of the effects of T in the sexual dimorphism of the CNS are exerted via estrogen receptor after their local aromatization to estradiol.

To our best knowledge, our study provides the first evidence that the expression of both 5-R isozymes in the brain, at least in the cerebral cortex, is regulated in opposite ways by androgens. This finding suggests that both isozymes may play a role in the sexual dimorphism of the CNS, besides other functions. Given that both isozymes are expressed and regulated in the adult animal, it appears that they participate not only in the formation and development of the brain, but also in the maintenance of sexually dimorphic brain structures, offering a dynamic rather than a static model of the cerebral functionality required by the physiology and psychology of the individual.

	ACKNOWLEDGMENTS

 
We thank R. Davies for revising the English text and the Clinico University Hospital of Granada for letting us use the ABIPRISM 310 Genetic Analyzer. This work was founded in part by DGICYT PM97-0177 and the Andalusian Regional Government (Endocrinology and Metabolism Group).

Received for publication November 20, 2002. Accepted for publication March 28, 2003.

	REFERENCES

TOP ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 

1. Poletti, A., Negri-Cesi, P., Rabuffetti, M., Colciago, A., Celotti, F., Martini, L. (1998) Transient expression of the 5alpha-reductase type 2 isozyme in the rat brain in late fetal and early postnatal life. Endocrinology 139,2171-2178[Abstract/Free Full Text]
2. Poletti, A., Celotti, F., Rumio, C., Rabuffetti, M., Martini, L. (1997) Identification of type 1 5alpha-reductase in myelin membranes of male and female rat brain. Mol. Cell. Endocrinol. 129,181-190[CrossRef][Medline]
3. Melcangi, R. C., Poletti, A., Cavarretta, I., Celotti, F., Colciago, A., Magnaghi, V., Motta, M., Negri-Cesi, P., Martini, L. (1998) The 5-reductase in the central nervous system: expression and modes of control. J. Steroid Biochem. Mol. Biol. 65,295-299[CrossRef][Medline]
4. Lauber, M. E., Lichtensteiger, W. (1996) Ontogeny of 5a-reductase (Type1) messenger ribonucleic acid expression in rat brain early presence in germinal zones. Endocrinology 137,2718-2730[Abstract]
5. Mellon, S. H., Griffin, L. D. (2002) Neurosteroids: biochemistry and clinical significance. Trends Endocrinol. Metab. 13,35-43[CrossRef][Medline]
6. Barbaccia, M. L., Serra, M., Purdy, R. H., Biggio, G. (2001) Stress and neuroactive steroids. Int. Rev. Neurobiol. 46,243-272[Medline]
7. Purdy, R. H., Morrow, A. L., Moore, P. H., Jr, Paul, S. M. (1991) Stress-induced elevations of gamma-aminobutyric acid type A receptor-active steroids in the rat brain. Proc. Natl. Acad. Sci. USA 88,4553-4557[Abstract/Free Full Text]
8. Barbaccia, M. L., Roscetti, G., Trabucchi, M., Mostallino, M. C., Concas, A., Purdy, R. H., Biggio, G. (1996) Time-dependent changes in rat brain neuroactive steroid concentrations and GABAA receptor function after acute stress. Neuroendocrinology 63,166-172[Medline]
9. Barbaccia, M. L., Roscetti, G., Trabucchi, M., Purdy, R. H., Mostallino, M. C., Concas, A., Biggio, G. (1997) The effects of inhibitors of GABAergic transmission and stress on brain and plasma allopregnanolone concentrations. Br. J. Pharmacol. 120,1582-1588[CrossRef][Medline]
10. Finn, D. A., Gee, K. W. (1994) The estrus cycle, sensitivity to convulsants and the anticonvulsant effect of a neuroactive steroid. J. Pharmacol. Exp. Ther. 271,164-170[Abstract/Free Full Text]
11. Concas, A., Mostallino, M. C., Porcu, P., Follesa, P., Barbaccia, M. L., Trabucchi, M., Purdy, R. H., Grisenti, P., Biggio, G. (1998) Role of brain allopregnanolone in the plasticity of gamma-aminobutyric acid type A receptor in rat brain during pregnancy and after delivery. Proc. Natl. Acad. Sci. USA 95,13284-13289[Abstract/Free Full Text]
12. Biggio, G., Barbaccia, M. L., Follesa, P., Serra, M., Purdy, R. H., Concas, A. (1999) Neurosteroids and GABA_A receptor plasticity. Martin, D. L. Olsen, R. W. eds. GABA in the Nervous System Lippincott-Williams and Wilkins
13. Celotti, F., Melcangi, R. C., Martini, L. (1992) The 5 alpha-reductase in the brain: molecular aspects and relation to brain function. Front. Neuroendocrinol. 13,163-215[Medline]
14. Lambert, J. J., Belelli, D., Hill-Venning, C., Peters, J. A. (1995) Neurosteroids and GABAA receptor function. Trends Pharmacol. Sci. 16,295-303[CrossRef][Medline]
15. Torres, J. M., Ortega, E. (2002) Alcohol intoxication increases allopregnanolone levels in female adolescents human. Neuropsychopharmacology In press
16. Serra, M., Pisu, M. G., Littera, M., Papi, G., Sanna, E., Tuveri, F., Usala, L., Purdy, R. H., Biggio, G. (2000) Social isolation-induced decreases in both the abundance of neuroactive steroids and GABA(A) receptor function in rat brain. J. Neurochem. 75,732-740[CrossRef][Medline]
17. Paul, S. M., Purdy, R. H. (1992) Neuroactive steroids. FASEB J. 6,2311-2322[Abstract]
18. Compagnone, N. A., Mellon, S. H. (2000) Neurosteroids: biosynthesis and function of these novel neuromodulators. Front. Neuroendocrinol. 21,1-56[CrossRef][Medline]
19. George, F. W., Russell, D. W., Wilson, J. D. (1991) Feed-forward control of prostate growth: dihydrotestosterone induces expression of its own biosynthetic enzyme, steroid 5 alpha-reductase. Proc. Natl. Acad. Sci. USA 88,8044-8047[Abstract/Free Full Text]
20. Torres, J. M., Gomez-Capilla, J. A., Ortega, E. (2002) Quantitative reverse-transcriptase-polymerase chain reaction assay for mRNA levels of steroid 5-reductase isozymes. Anal. Biochem. 307,177-180[CrossRef][Medline]
21. Chomczynski, P., Sacchi, N. (1987) Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal. Biochem. 162,156-159[Medline]
22. Negri-Cesi, P., Poletti, A., Celotti, F. (1996) Metabolism of steroids in the brain: a new insight into the role of 5alpha-reductase and aromatase in brain differentiation and functions. J. Steroid Biochem. Mol. Biol. 58,455-466[CrossRef][Medline]
23. Russell, D. W., Wilson, J. D. (1994) Steroid 5-reductase: two genes/two enzymes. Annu. Rev. Biochem. 63,25-61[Medline]
24. Torres, J. M., Ruiz, E., Ortega, E. (2002) Development of a quantitative RT-PCR method to study 5-reductase mRNA isozymes in rat prostate in different androgen status. Prostate In press
25. Poletti, A., Coscarella, A., Negri-Cesi, P., Colciago, A., Celotti, F., Martini, L. (1998) 5 alpha-reductase isozymes in the central nervous system. Steroids 63,246-251[CrossRef][Medline]
26. Mahendroo, M. S., Cala, K. M., Landrum, D. P., Russell, D. W. (1997) Fetal death in mice lacking 5alpha-reductase type 1 caused by estrogen excess. Mol. Endocrinol. 11,917-927[Abstract/Free Full Text]
27. Normington, K., Russell, D. W. (1992) Tissue distribution and kinetic characteristics of rat steroid 5-reductase isozymes. J. Biol. Chem. 267,19548-19554[Abstract/Free Full Text]
28. Torres, J. M., Ruiz, E., Ortega, E. (2001) Effects of CRH and ACTH administration on plasma and brain neurosteroid levels. Neurochem. Res. 26,555-558[CrossRef][Medline]
29. Corpechot, C., Young, J., Calvel, M., Wehrey, C., Veltz, J. N., Touyer, G., Mouren, M., Prasad, V. V., Banner, C., Sjovall, J., et al (1993) Neurosteroids: 3 alpha-hydroxy-5 alpha-pregnan-20-one and its precursors in the brain, plasma, and steroidogenic glands of male and female rats. Endocrinology 133,1003-1009[Abstract]
30. Segovia, S., Guillamon, A., del Cerro, M. C. R., Ortega, E., Perez-Laso, C., Rodriguez, M., Beyer, C. (1999) The development of brain sex differences: a multisignaling process. Behav. Brain Res. 105,69-80[CrossRef][Medline]
31. Segovia, S., del Cerro, M. C. R., Ortega, E., Perez-Laso, C., Rodriguez, M., Izquierdo, M. A. P., Gillamon, A. (1996) Role of GABA A receptors in the organization of brain and behavioural sex differences. NeuroReport 7,2553-2557[Medline]
32. Majewska, M. D. (1992) Neurosteroids: endogenous bimodal modulators of the GABAA receptor. Mechanism of action and physiological significance. Prog. Neurobiol. 38,379-395[CrossRef][Medline]
33. Li, X., Bertics, P. J., Karavolas, H. J. (1997) Regional distribution of cytosolic and particulate 5alpha-dihydroprogesterone 3 alpha-hydroxysteroid oxidoreductases in female rat brain. J. Steroid Biochem. Mol. Biol. 60,311-318[CrossRef][Medline]
34. Einarsson, K., Gustafsson, J. A., Stenberg, A. (1973) Neonatal imprinting of liver microsomal hydroxilation and reduction steroids. J. Biol. Chem. 248,4987-4997[Abstract/Free Full Text]
35. Valencia, A., Collado, P., Cales, J. M., Segovia, S., Perez-Laso, C., Rodríguez-Zafra, M., Guillamon, A. (1992) Postnatal administration of dihydrotestosterone to the male rat abolishes sexual dimorphism in the accessory olfactory bulb: a volumetric study. Brain Res. Dev. Brain Res. 8,132-135
36. Matsui, D., Sakari, M., Sato, T., Murayama, A., Takada, I., Kim, M., Takeyama, K., Kato, S. (2002) Transcriptional regulation of the mouse steroid 5-alpha-reductase type II gene by progesterone in brain. Nucleic Acids Res. 30,1387-1393[Abstract/Free Full Text]
37. Lephart, E. D. (1996) A review of brain aromatase cytochrome P450. Brain Res. 22,1-26

This article has been cited by other articles:

				N. A. Henderson, G. M Cooke, and B. Robaire Region-specific expression of androgen and growth factor pathway genes in the rat epididymis and the effects of dual 5{alpha}-reductase inhibition. J. Endocrinol., September 1, 2006; 190(3): 779 - 791. [Abstract] [Full Text] [PDF]

				M. SCHMIDT, H. NAUMANN, C. WEIDLER, M. SCHELLENBERG, S. ANDERS, and R. H. STRAUB Inflammation and Sex Hormone Metabolism Ann. N.Y. Acad. Sci., June 1, 2006; 1069(1): 236 - 246. [Abstract] [Full Text] [PDF]

				J M Torres and E Ortega Steroid 5{alpha}-reductase isozymes in the adult female rat brain: central role of dihydrotestosterone. J. Mol. Endocrinol., April 1, 2006; 36(2): 239 - 245. [Abstract] [Full Text] [PDF]

				S. Thiele, U. Hoppe, P.-M. Holterhus, and O. Hiort Isoenzyme type 1 of 5alpha-reductase is abundantly transcribed in normal human genital skin fibroblasts and may play an important role in masculinization of 5alpha-reductase type 2 deficient males Eur. J. Endocrinol., June 1, 2005; 152(6): 875 - 880. [Abstract] [Full Text] [PDF]

				G. Pinna, E. Costa, and A. Guidotti Changes in brain testosterone and allopregnanolone biosynthesis elicit aggressive behavior PNAS, February 8, 2005; 102(6): 2135 - 2140. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by TORRES, J. M. Articles by ORTEGA, E. Search for Related Content PubMed PubMed Citation Articles by TORRES, J. M. Articles by ORTEGA, E.