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Possible interaction of the adrenal-gonadal systems on brain catecholamines of adult male rats
Camp. Biochem. Physiol. Vol. 86C, No. 2, pp. 295-298, Printed in Great Britain
1987
0
0306-4492/87 $3.00+ 0.00 1987Pergamon Journals Ltd
POSSIBLE INTERACTION OF THE ADRENAL-GONADAL SYSTEMS ON BRAIN CATECHOLAMINES OF ADULT MALE RATS Departamento
M. L. LERET, P. TRANQLJE, I. GONZALEZ and J. C. CALVO de Fisiologia Animal, Facultad de Ciencias Biologicas, Universidad Complutense, Madrid 28040, Spain (Received 19 May 1986)
Abstract-l. Studies from this laboratory showed that gonadectomy (GDX) alters biogenic amines concentrations in diencephalon during the first 40 days. 2. While the GDX females maintain the differences at day 60, the differences are eliminated in males at that time. 3. In the present work, we have studied in three cerebral regions the adrenal involvement in the mechanism responsible for this normalization of catecholamine concentration in long-term castrated adult male rats. 4. A hypersecretion of adrenal steroids seems to compensate for the lack of gonadal effect when the orchidectomized rats reach adulthood only for diencephalic dopamine.
INTRODUCTION
these cells are directly affected by glucocorticoids or whether the adrenal steroids produce their effects via an indirect trans-synaptic mechanism. Diverse studies suggest that plasma glucocorticoids have a possible effect on the metabolism of central NA (Kawa et al., 1977). A trans-synaptic modulation of noradrenergic cells by glucocorticoid target neurons is likely given the widespread distribution of the glucocorticoid sensitive cells in the CNS (Duncan and Stumpf, 1985). Among the anatomical regions which contain 3H-corticosterone target neurons are the hypothalamus, hippocampus, amygdala and lateral septal nuclei. Likewise, dopaminergic activity in the medio-basal hypothalamus is probably subject to modulation by corticosterone (Versteeg et al., 1984). The effect of plasma glucocorticoids on central catecholamines is not necessarily related to the feedback mechanism of pituitary adrenocortical functions. Especially, involvement of the adrenal glands in the maturation and function of the rat brain pituitary-gonadal axis has been suggested in previous studies (Cartes-Gallegos et al., 1984). On the other hand, administration of testosterone and estradiol to intact rats indicates that both gondal hormones exert multiple effects on the pituitary-adrenal axis, acting both on adrenal gland level and on ACTH secretion (Kitay, 1963). Studies from this laboratory showed that gonadectomy (GDX) alters DA, NA and serotonin concentrations in diencephalon, and homovanillic acid levels in diencephalon, striatum and limbic system during the first 40 days following surgery (Leret and Fraile, 1985; Leret et al., 1984; Martinez-Conde et al., 1985). In adult female rats-60 days old-the differences between gonadectomized and intact animals were maintained. However, no differences were observed in adult male rats of the same age. We propose that the result found in the castrated 60-day-old males could be explained by a steroid hormones compensat-
The release of gonadotropin from the pituitary gland is under the modulatory control of central catecholaminergic neurons (Barraclough et al., 1984). It is generally accepted that regulation of gonadotropin secretion is maintained at least in part by feedback inhibition of androgenic steroids on the central nervous system (CNS). Thus, negative feedback by testosterone or its active metabolite 5-a-dihydrotestosterone suppresses luteinizing hormone releasing hormone (LHRH)-mediated LH release (Tibolt and Childs, 1985). Testosterone probably exerts its negative feedback effects at both hypothalamic and pituitary levels. At the former site, it has been shown to prevent the reduction in LHRH concentration in the medial basal hypothalamus which normally occurs after castration (Kalra and Kalra, 1978). A neurotransmitter mediator system may exist between the androgen-sensitive cells of the CNS and the releasing hormone producing neurons of the hypothalamus. Particularly, the catecholaminergic control of gonadotropin secretion seems to be involved in the feedback action of gonadal steroids (Ojeda and McCann, 1973; Kizer et al., 1978; Vermes et al., 1979; Simpkins et al., 1980). Major efforts have been directed towards studies of the feedback mechanisms of androgenic steroid within the hypothalamus, but recent studies suggest the limbic structures as additional loci of steroid feedback action in males (Vermes et al., 1979). On the other hand, there is considerable evidence to indicate that glucocorticoid hormones from the adrenal gland can modulate enzymatic processes involved in the synthesis of catecholamines in CNS. Iuvone et al. (1977) found that corticosterone treatment increases the conversion of systemically administered ‘H-tyrosine to ‘H-norepinephrine in endocrinologically intact mice. It is not known whether 295
296
M. L. LERETet al.
Fig. 1. Diencephalic (D), striatal (C.S.) and limbic (L.S.) DA levels (in pg/g of fresh tissue) in male rats after ORDX (B). ADX (m) or ORDX/ADX (m) compared to sham-controls (0). All the animals were
operated at day 21 and sacrificed at day 60 of age. Values represent means k SEM. (*P < 0.05, **p < 0.025, ***p i 0.01, ****p < 0.005.) ory hypersecretion caused by the absence of testicular androgens. The present experiments were performed to investigate if the adrenal gland is in charge of carrying out this compensation. For this purpose, the animals were orchidectomized (ORDX), adrenalectomized (ADX) and ORDX +ADX at weaning day. Diencephalic, striatal and limbic DA and NA were measured at adulthood, to determine the adrenal involvement in the hypothetically compensatory effect previously mentioned. MATERIALS
AND METHODS
Albino male rats of a laboratory inbred SpragueeDawley strain were used throughout. Animals were fed ad libitum and kept under normal daylight conditions at a constant temperature (21 * 2°C). The pups were weaned at 21 days after birth. Orchidectomy (ORDX), adrenalectomy (ADX), ORDX plus ADX or sham operations were performed at day 21: all operative procedures were carried out under ether anaesthesia and ADX animals were given a 0.9% sodium chloride solution postoperatively. The animals were housed under constant conditions till the time of sacrifice. At 60 days after birth, all the groups were sacrificed by decapitation between 9 and II a.m., in order to avoid the catecholamine circadian rhythm.
The brains were rapidly removed. The diencephalon, corpus striatum and limbic system were dissected, cut according to Carlsson and Lindqvist (1973) on a cold surface. Brain tissue samples were weighed and stored at -20°C until analysis for not more than 2-3 weeks, because after this time it was observed that important catecholamine losses occurred. Dopamine (DA) and noradrenaline (NA) were extracted and assayed using a method adopted from Welch and Welch (1969). A Perkin-Elmer model 150 spectrophotofluorometer was used. Statistical analysis Data are expressed as mean +_SEM. Differences among groups in DA and NA content of every brain area were tested using analysis of variance followed by a Scheffe test. In the figures, only the significance between the control group and every other group is shown. RESULTS
DA levels in diencephalon
Figure 1 shows that ORDX did not significantly alter DA levels in diencephalon on day 60 postcastration. In contrast, ADX-ORDX produced an acute increase compared with sham control, orchidectomized and adrenalectomized groups.
Fig. 2. Graph showing the variations of diencephalic (D), striatal (C.S.) and limbic (L.S.) NA levels (in pg/g of fresh tissue) of 60-day-old male rats ADX (@), ORDX (m) or ADXjORDX (am) operated at day 21, compared to sham controls (0). Results are expressed as arithmetic mean & SEM. (*P < 0.05, **p < 0.025, ***p < 0.01, ****p < 0.005.)
Gonadectomy,
adrenalectomy and catecholamines
DA level in striatum Corpus striatum concentrations of DA were not significantly changed in any of the groups compared with sham controls. The only significant difference was observed between adrenalectomized and adrenalectomized-orchidectomized animals (see Fig. 1). DA level in limbic system In the limbic system, we found an acute increase in orchidectomized and adrenalectomized-orchidectomized rats, while ADX kept limbic DA levels close to the control ones (see Fig. 1). NA levels in diencephalon In diencephalon, the NA concentration was raised in ADX-ORDX animals above the ADX and ORDX values, but this change was not significant (see Fig. 2). NA levels in striatum ADX, ORDX and double surgery enhanced NA levels in the striatum with respect to the control group. NA levels reached similar values in these groups. Significant differences were only found between the ADX-ORDX group and controls (see Fig. 2). NA levels in limbic system Figure 2 shows that no changes were observed in the limbic level of NA after the different endocrine manipulations. DISCUSSION
In the experiments described so far, we found no correlation between the effects of castration and alteration in the hypothalamic catecholaminergic system. Similar results have been reported in previous works. Thus, no significant changes were found both in NA concentration at different intervals after castration (Vermes et al., 1979; Herdon et al., 1984) and DA concentration 4 weeks after ORDX (Vermes et al., 1979), in whole hypothalamus. In the median eminence, Kizer et al. (1978) and Lofstrom (1979, 1980) found no alteration in NA level 10 and 30 days after castration. In contrast, some studies in discrete hypothalamic areas have reported that NA concentration increases in the anterior hypothalamus and median eminence within 8 hr of castration (Chiocchio et al., 1976; de Paolo et al., 1982) and that NA concentration in the anterior hypothalamus is increased 10-20 days after castration (Donoso et al., 1967). On the other hand, Vermes et al. (1979) did not find alteration of DA content either in the anterior part of the hypothalamus or in the median eminence, following gonadectomy. The discrepancies among the above described results may be partially due to the different extent of neural tissue examined for DA and NA analysis. Moreover, different postcastrational intervals have been chosen in each particular experiment. Thus, in previous studies performed in this laboratory, a high increase in the diencephalic DA and NA levels 7 and 22 days after castration was observed. However, in 60-day-old rats (37 days postcastration). DA values were similar to those of sham controls and NA
291
concentration was, in turn, hardly higher (Leret and Fraile, 1985). In addition, a sex difference has been seen in the response of the hypothalamic NA and DA systems to gonadectomy. Thus we did not find changes in DA and NA levels 37 days after ORDX. Conversely, previous works showed that ovariectomy results in a significant enhancement in catecholamine values with respect to sham controls (Leret and Fraile, 1985). In agreement with these data, Herdon et al. (1984) found that phenoxybenzamine, an alpha-adrenergic antagonist, did not alter LH levels on day 40 after gonadectomy only in male rats. We performed ADX in order to check the possible participation of the adrenal glands in the above described sexual difference. The ADX-ORDX group showed significantly greater catecholamine levels in diencephalon with respect to only the castrated and the intact animals. This result suggests that the lack of differences between the control and the ORDX group may be due to an adrenal hypersecretion in the 60-days castrated male rats. Therefore, the adrenal steroid secretion would substitute the gonadal steroid secretion in such conditions. The mechanism involved in the increase of the adrenal secretion 37 days after ORDX remains unknown. Nevertheless, it seems to be associated with the sexual maturation of the male rat. On the other hand, our results support the involvement of DA from limbic system structures in the central gonadal steroid feedback action, whereas neither the gonadal nor the adrenal steroids affected the NA content of the limbic system. Previous works considered the limbic structures as additional loci of steroid feedback action (McEwen et al., 1970), and Vermes et al. (1979) found that administration of testosterone propionate (5 pg/kg) decreased the DA level in the amygdala to a minimum after 90-120 min. However, this dose was ineffective as regards to altering NA levels in this limbic stricture. Shen and Ganong (1976) observed no changes in the NA and DA concentration of the hippocampus after ADX. This dose was ineffective as regards altering NA levels in this limbic structure. Shen and Ganong observed no changes in the NA or DA concentration of the hippocampus after ADX. It has been suggested that the corpus striatum could be another extrahypothalamic target structure for hormone action (Ramirez, 1983). However, our findings are in contrast to this proposal, for no remarkable changes in NA and DA concentrations were observed after ORDX or ADX. (Only ORDX and ADX together resulted in a light increase in NA levels.) Vermes et al. (1979) obtained somewhat similar results to ours measuring catecholamine levels for the first 120 min after testosterone propionate treatment.
REFERENCES
Barraclough C. A., Wise P. M. and Selmanoff
M. K. (1984) A rBle for hypothalamic catecholamines in the regulation of gonadotropin secretion. Rec. Prog. Horm. Res. 40, 487-592. Carlsson A. and Lindqvist M. (1973) Effect of ethanol on hydroxylation of tyrosine and trytophan in rat brain in
vivo. J. Pharm. Phnrmac. 25, 437440.
298
M. L. LERETet al.
Chiocchio S. R., Negro-Vilar A. and Tramezzani J. H. (1976) Acute changes in norepinephrine content in the median eminence induced by orchidectomy or testosterone replacement, Endocrinology 99, 629dj5. Cartes-Gallegos V., Alonso R., Castafieda I;., Sojo I., Carranco A., Cervantes C. and Parra A. (1984) Paramethasone acetate (PA): corticosteroid potency vs hypothalamic pituitary-gonadal axis. J. Steroid Biochem. 20, 353-356. Donoso A. O., Stefano F. J. E., Biscardi A. M. and Cukier J. (1967) Effects of castration on hypothalamic catecholamines. Am. J. Physiol. 212, 737-739. Duncan G. E. and Stumpf W. E. (1985) A combined autoradiographic and immunocytochemical study of 3Hcorticosterone target neurons and catecholamine neurons in rat and mouse lower brain stem. Neuroendocrinology 40, 262-27 1. Herdon H. J.. Everard D. M. and Wilson C. A. (1984) Studies on the control of gonadotrophin release rn the gonadectomized male rat: evidence for a lack of involvement of the hypothalamic noradrenergic system in the long-term castrated rat. J. Endocr. 100, 235-244. Iuvone P. M., Morasco J. and Dunn A. J. (1977) Effect of corticosterone on the synthesis of 3H-catecholamines in the brains of CD-l mice. Brain Res. 120, 571-576. Kalra P. S. and Kalra S. P. (1978) Effects of intrahypothalamic testosterone implants on LHRH levels in the preoptic area and the medial basal hypothalamus. Life Sci. 23, 65-68. Kawa A., Kamisaki T., Ariyama T., Maeda Y. and Kanehisa T. (1977) The effects of adrenalectomy and the administration of dexamethasone on the levels and turnover rate of brain noradrenaline in rats. IRCS Med. Sci. 5, 449. Kitay J. I. (1963) Pituitary-adrenal function in the rat after gonadectomy and gonadal hormone replacement. Endocrinology 73, 253-260. Kizer J. S., Humm J., Nicholson G., Greely G. and Youngblood W. (1978) The effect of castration, thyroidectomy and haloperidol upon the turnover rates of dopamine and norepinephrine and the kinetic properties of tyrosine hydroxylase in discrete hypothalamic nuclei of the male rat. Brain Res. 146, 955107. Leret M. L. and Fraile A. (1985) Effect of gonadectomy on brain catecholamines during the postnatal period. Comp. Biochem. Physiol. 81C, 405409. Leret M. L., Olid J. M. and Martinez-Conde E. (1984) Effect of gonadectomy on brain homovanillic acid levels. Comp. Biochem. Physiol. 78B, 773-776.
Lofstrom A. (1979) Catecholamine content of the rat median eminence following removal of endocrine glands. Psvchoneuroendocrinology 4, 57-65. Lofstrom A. (1980) Increase in medial palisade zone dopamine activity after long-term castration in the male rat. Endokrinologie 76, 23-28. Martinez-Conde E., Leret M. L. and Diaz S. (1985) The influence of testosterone in the brain of the male rat on levels of serotonin (5-HT) and hydroxyindoleacetic acid (5-HIAA). Comp. Biochem. Physiol. 8OC, 41 l-414. McEwen B. S., Pfaff D. W. and Zigmon R. E. (1970) Factors influencing sex hormone uptake by rat brain regions-III. Effects of competing steroid on testosterone uptake. Brain Res. 21, 29-38. Ojeda S. R. and McCann S. M. (1973) Evidence for participation of a catecholaminergic mechanism in the post-castration rise in plasma gonadotropins. NeuroendocrinoloEv 12. 295-3 15. de Paolo-L. V:, McCann S. M. and Negro-Vilar A. (1982) A sex difference in the activation of hypothalamic catecholaminergic and LHRH peptidergic neurons after acute castration. Endocrinology 110, 531-539. Ramirez V. D. (1983) Hormones and striatal dopaminergic activity: a novel neuroendocrine model. In The Anterior Pituitary Gland (Edited by Ajay S. and Bhatnagar), pp. 97-105. Raven Press, New York. Shen J. T. and Ganong W. F. (1976) Effect of Variations in adrenocortical function on dopamine b-hydroxylase and norepinephrine in the brain of the rat. J. Pharmac. exp. Ther. 199, 639448. Simpkins J. W., Kalra P. S. and Kalra S. P. (1980) Effects of testosterone on cathecholamine turnover and LHRH content in the basal hypthalamus and preoptic area. Neuroendocrinology 30, 94100. Tibolt R. E. and Childs G. V. (1985) Cytochemical and cytophysiological studies of gonadotropin releasing hormone (GnRH) target cells in the male rat pituitary: differential effects of androgens and corticosterone on GnRH binding and gonadotropin release. Endocrinology 117, 396-404. Vermes I., Varszegi M., Totti E. K. and Telegdy G. (1979) Action of androgenic steroids on brain neurotransmitters in rats. Neuroendocrinology 28, 386393. Versteed D. H. G., Van Zoest I. and De Kloet E. R. (1984) Acute changes in dopamine metabolism in the media basal hypothalamus following adrenalectomy. Experientia 40, 112-114. Welch A. and Welch B. L. (1969) NE, DA, 5-HT and 5-HTAA in mouse brain. Analyt. Biochem. 30, 161-179.
1987
0
0306-4492/87 $3.00+ 0.00 1987Pergamon Journals Ltd
POSSIBLE INTERACTION OF THE ADRENAL-GONADAL SYSTEMS ON BRAIN CATECHOLAMINES OF ADULT MALE RATS Departamento
M. L. LERET, P. TRANQLJE, I. GONZALEZ and J. C. CALVO de Fisiologia Animal, Facultad de Ciencias Biologicas, Universidad Complutense, Madrid 28040, Spain (Received 19 May 1986)
Abstract-l. Studies from this laboratory showed that gonadectomy (GDX) alters biogenic amines concentrations in diencephalon during the first 40 days. 2. While the GDX females maintain the differences at day 60, the differences are eliminated in males at that time. 3. In the present work, we have studied in three cerebral regions the adrenal involvement in the mechanism responsible for this normalization of catecholamine concentration in long-term castrated adult male rats. 4. A hypersecretion of adrenal steroids seems to compensate for the lack of gonadal effect when the orchidectomized rats reach adulthood only for diencephalic dopamine.
INTRODUCTION
these cells are directly affected by glucocorticoids or whether the adrenal steroids produce their effects via an indirect trans-synaptic mechanism. Diverse studies suggest that plasma glucocorticoids have a possible effect on the metabolism of central NA (Kawa et al., 1977). A trans-synaptic modulation of noradrenergic cells by glucocorticoid target neurons is likely given the widespread distribution of the glucocorticoid sensitive cells in the CNS (Duncan and Stumpf, 1985). Among the anatomical regions which contain 3H-corticosterone target neurons are the hypothalamus, hippocampus, amygdala and lateral septal nuclei. Likewise, dopaminergic activity in the medio-basal hypothalamus is probably subject to modulation by corticosterone (Versteeg et al., 1984). The effect of plasma glucocorticoids on central catecholamines is not necessarily related to the feedback mechanism of pituitary adrenocortical functions. Especially, involvement of the adrenal glands in the maturation and function of the rat brain pituitary-gonadal axis has been suggested in previous studies (Cartes-Gallegos et al., 1984). On the other hand, administration of testosterone and estradiol to intact rats indicates that both gondal hormones exert multiple effects on the pituitary-adrenal axis, acting both on adrenal gland level and on ACTH secretion (Kitay, 1963). Studies from this laboratory showed that gonadectomy (GDX) alters DA, NA and serotonin concentrations in diencephalon, and homovanillic acid levels in diencephalon, striatum and limbic system during the first 40 days following surgery (Leret and Fraile, 1985; Leret et al., 1984; Martinez-Conde et al., 1985). In adult female rats-60 days old-the differences between gonadectomized and intact animals were maintained. However, no differences were observed in adult male rats of the same age. We propose that the result found in the castrated 60-day-old males could be explained by a steroid hormones compensat-
The release of gonadotropin from the pituitary gland is under the modulatory control of central catecholaminergic neurons (Barraclough et al., 1984). It is generally accepted that regulation of gonadotropin secretion is maintained at least in part by feedback inhibition of androgenic steroids on the central nervous system (CNS). Thus, negative feedback by testosterone or its active metabolite 5-a-dihydrotestosterone suppresses luteinizing hormone releasing hormone (LHRH)-mediated LH release (Tibolt and Childs, 1985). Testosterone probably exerts its negative feedback effects at both hypothalamic and pituitary levels. At the former site, it has been shown to prevent the reduction in LHRH concentration in the medial basal hypothalamus which normally occurs after castration (Kalra and Kalra, 1978). A neurotransmitter mediator system may exist between the androgen-sensitive cells of the CNS and the releasing hormone producing neurons of the hypothalamus. Particularly, the catecholaminergic control of gonadotropin secretion seems to be involved in the feedback action of gonadal steroids (Ojeda and McCann, 1973; Kizer et al., 1978; Vermes et al., 1979; Simpkins et al., 1980). Major efforts have been directed towards studies of the feedback mechanisms of androgenic steroid within the hypothalamus, but recent studies suggest the limbic structures as additional loci of steroid feedback action in males (Vermes et al., 1979). On the other hand, there is considerable evidence to indicate that glucocorticoid hormones from the adrenal gland can modulate enzymatic processes involved in the synthesis of catecholamines in CNS. Iuvone et al. (1977) found that corticosterone treatment increases the conversion of systemically administered ‘H-tyrosine to ‘H-norepinephrine in endocrinologically intact mice. It is not known whether 295
296
M. L. LERETet al.
Fig. 1. Diencephalic (D), striatal (C.S.) and limbic (L.S.) DA levels (in pg/g of fresh tissue) in male rats after ORDX (B). ADX (m) or ORDX/ADX (m) compared to sham-controls (0). All the animals were
operated at day 21 and sacrificed at day 60 of age. Values represent means k SEM. (*P < 0.05, **p < 0.025, ***p i 0.01, ****p < 0.005.) ory hypersecretion caused by the absence of testicular androgens. The present experiments were performed to investigate if the adrenal gland is in charge of carrying out this compensation. For this purpose, the animals were orchidectomized (ORDX), adrenalectomized (ADX) and ORDX +ADX at weaning day. Diencephalic, striatal and limbic DA and NA were measured at adulthood, to determine the adrenal involvement in the hypothetically compensatory effect previously mentioned. MATERIALS
AND METHODS
Albino male rats of a laboratory inbred SpragueeDawley strain were used throughout. Animals were fed ad libitum and kept under normal daylight conditions at a constant temperature (21 * 2°C). The pups were weaned at 21 days after birth. Orchidectomy (ORDX), adrenalectomy (ADX), ORDX plus ADX or sham operations were performed at day 21: all operative procedures were carried out under ether anaesthesia and ADX animals were given a 0.9% sodium chloride solution postoperatively. The animals were housed under constant conditions till the time of sacrifice. At 60 days after birth, all the groups were sacrificed by decapitation between 9 and II a.m., in order to avoid the catecholamine circadian rhythm.
The brains were rapidly removed. The diencephalon, corpus striatum and limbic system were dissected, cut according to Carlsson and Lindqvist (1973) on a cold surface. Brain tissue samples were weighed and stored at -20°C until analysis for not more than 2-3 weeks, because after this time it was observed that important catecholamine losses occurred. Dopamine (DA) and noradrenaline (NA) were extracted and assayed using a method adopted from Welch and Welch (1969). A Perkin-Elmer model 150 spectrophotofluorometer was used. Statistical analysis Data are expressed as mean +_SEM. Differences among groups in DA and NA content of every brain area were tested using analysis of variance followed by a Scheffe test. In the figures, only the significance between the control group and every other group is shown. RESULTS
DA levels in diencephalon
Figure 1 shows that ORDX did not significantly alter DA levels in diencephalon on day 60 postcastration. In contrast, ADX-ORDX produced an acute increase compared with sham control, orchidectomized and adrenalectomized groups.
Fig. 2. Graph showing the variations of diencephalic (D), striatal (C.S.) and limbic (L.S.) NA levels (in pg/g of fresh tissue) of 60-day-old male rats ADX (@), ORDX (m) or ADXjORDX (am) operated at day 21, compared to sham controls (0). Results are expressed as arithmetic mean & SEM. (*P < 0.05, **p < 0.025, ***p < 0.01, ****p < 0.005.)
Gonadectomy,
adrenalectomy and catecholamines
DA level in striatum Corpus striatum concentrations of DA were not significantly changed in any of the groups compared with sham controls. The only significant difference was observed between adrenalectomized and adrenalectomized-orchidectomized animals (see Fig. 1). DA level in limbic system In the limbic system, we found an acute increase in orchidectomized and adrenalectomized-orchidectomized rats, while ADX kept limbic DA levels close to the control ones (see Fig. 1). NA levels in diencephalon In diencephalon, the NA concentration was raised in ADX-ORDX animals above the ADX and ORDX values, but this change was not significant (see Fig. 2). NA levels in striatum ADX, ORDX and double surgery enhanced NA levels in the striatum with respect to the control group. NA levels reached similar values in these groups. Significant differences were only found between the ADX-ORDX group and controls (see Fig. 2). NA levels in limbic system Figure 2 shows that no changes were observed in the limbic level of NA after the different endocrine manipulations. DISCUSSION
In the experiments described so far, we found no correlation between the effects of castration and alteration in the hypothalamic catecholaminergic system. Similar results have been reported in previous works. Thus, no significant changes were found both in NA concentration at different intervals after castration (Vermes et al., 1979; Herdon et al., 1984) and DA concentration 4 weeks after ORDX (Vermes et al., 1979), in whole hypothalamus. In the median eminence, Kizer et al. (1978) and Lofstrom (1979, 1980) found no alteration in NA level 10 and 30 days after castration. In contrast, some studies in discrete hypothalamic areas have reported that NA concentration increases in the anterior hypothalamus and median eminence within 8 hr of castration (Chiocchio et al., 1976; de Paolo et al., 1982) and that NA concentration in the anterior hypothalamus is increased 10-20 days after castration (Donoso et al., 1967). On the other hand, Vermes et al. (1979) did not find alteration of DA content either in the anterior part of the hypothalamus or in the median eminence, following gonadectomy. The discrepancies among the above described results may be partially due to the different extent of neural tissue examined for DA and NA analysis. Moreover, different postcastrational intervals have been chosen in each particular experiment. Thus, in previous studies performed in this laboratory, a high increase in the diencephalic DA and NA levels 7 and 22 days after castration was observed. However, in 60-day-old rats (37 days postcastration). DA values were similar to those of sham controls and NA
291
concentration was, in turn, hardly higher (Leret and Fraile, 1985). In addition, a sex difference has been seen in the response of the hypothalamic NA and DA systems to gonadectomy. Thus we did not find changes in DA and NA levels 37 days after ORDX. Conversely, previous works showed that ovariectomy results in a significant enhancement in catecholamine values with respect to sham controls (Leret and Fraile, 1985). In agreement with these data, Herdon et al. (1984) found that phenoxybenzamine, an alpha-adrenergic antagonist, did not alter LH levels on day 40 after gonadectomy only in male rats. We performed ADX in order to check the possible participation of the adrenal glands in the above described sexual difference. The ADX-ORDX group showed significantly greater catecholamine levels in diencephalon with respect to only the castrated and the intact animals. This result suggests that the lack of differences between the control and the ORDX group may be due to an adrenal hypersecretion in the 60-days castrated male rats. Therefore, the adrenal steroid secretion would substitute the gonadal steroid secretion in such conditions. The mechanism involved in the increase of the adrenal secretion 37 days after ORDX remains unknown. Nevertheless, it seems to be associated with the sexual maturation of the male rat. On the other hand, our results support the involvement of DA from limbic system structures in the central gonadal steroid feedback action, whereas neither the gonadal nor the adrenal steroids affected the NA content of the limbic system. Previous works considered the limbic structures as additional loci of steroid feedback action (McEwen et al., 1970), and Vermes et al. (1979) found that administration of testosterone propionate (5 pg/kg) decreased the DA level in the amygdala to a minimum after 90-120 min. However, this dose was ineffective as regards to altering NA levels in this limbic stricture. Shen and Ganong (1976) observed no changes in the NA and DA concentration of the hippocampus after ADX. This dose was ineffective as regards altering NA levels in this limbic structure. Shen and Ganong observed no changes in the NA or DA concentration of the hippocampus after ADX. It has been suggested that the corpus striatum could be another extrahypothalamic target structure for hormone action (Ramirez, 1983). However, our findings are in contrast to this proposal, for no remarkable changes in NA and DA concentrations were observed after ORDX or ADX. (Only ORDX and ADX together resulted in a light increase in NA levels.) Vermes et al. (1979) obtained somewhat similar results to ours measuring catecholamine levels for the first 120 min after testosterone propionate treatment.
REFERENCES
Barraclough C. A., Wise P. M. and Selmanoff
M. K. (1984) A rBle for hypothalamic catecholamines in the regulation of gonadotropin secretion. Rec. Prog. Horm. Res. 40, 487-592. Carlsson A. and Lindqvist M. (1973) Effect of ethanol on hydroxylation of tyrosine and trytophan in rat brain in
vivo. J. Pharm. Phnrmac. 25, 437440.
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M. L. LERETet al.
Chiocchio S. R., Negro-Vilar A. and Tramezzani J. H. (1976) Acute changes in norepinephrine content in the median eminence induced by orchidectomy or testosterone replacement, Endocrinology 99, 629dj5. Cartes-Gallegos V., Alonso R., Castafieda I;., Sojo I., Carranco A., Cervantes C. and Parra A. (1984) Paramethasone acetate (PA): corticosteroid potency vs hypothalamic pituitary-gonadal axis. J. Steroid Biochem. 20, 353-356. Donoso A. O., Stefano F. J. E., Biscardi A. M. and Cukier J. (1967) Effects of castration on hypothalamic catecholamines. Am. J. Physiol. 212, 737-739. Duncan G. E. and Stumpf W. E. (1985) A combined autoradiographic and immunocytochemical study of 3Hcorticosterone target neurons and catecholamine neurons in rat and mouse lower brain stem. Neuroendocrinology 40, 262-27 1. Herdon H. J.. Everard D. M. and Wilson C. A. (1984) Studies on the control of gonadotrophin release rn the gonadectomized male rat: evidence for a lack of involvement of the hypothalamic noradrenergic system in the long-term castrated rat. J. Endocr. 100, 235-244. Iuvone P. M., Morasco J. and Dunn A. J. (1977) Effect of corticosterone on the synthesis of 3H-catecholamines in the brains of CD-l mice. Brain Res. 120, 571-576. Kalra P. S. and Kalra S. P. (1978) Effects of intrahypothalamic testosterone implants on LHRH levels in the preoptic area and the medial basal hypothalamus. Life Sci. 23, 65-68. Kawa A., Kamisaki T., Ariyama T., Maeda Y. and Kanehisa T. (1977) The effects of adrenalectomy and the administration of dexamethasone on the levels and turnover rate of brain noradrenaline in rats. IRCS Med. Sci. 5, 449. Kitay J. I. (1963) Pituitary-adrenal function in the rat after gonadectomy and gonadal hormone replacement. Endocrinology 73, 253-260. Kizer J. S., Humm J., Nicholson G., Greely G. and Youngblood W. (1978) The effect of castration, thyroidectomy and haloperidol upon the turnover rates of dopamine and norepinephrine and the kinetic properties of tyrosine hydroxylase in discrete hypothalamic nuclei of the male rat. Brain Res. 146, 955107. Leret M. L. and Fraile A. (1985) Effect of gonadectomy on brain catecholamines during the postnatal period. Comp. Biochem. Physiol. 81C, 405409. Leret M. L., Olid J. M. and Martinez-Conde E. (1984) Effect of gonadectomy on brain homovanillic acid levels. Comp. Biochem. Physiol. 78B, 773-776.
Lofstrom A. (1979) Catecholamine content of the rat median eminence following removal of endocrine glands. Psvchoneuroendocrinology 4, 57-65. Lofstrom A. (1980) Increase in medial palisade zone dopamine activity after long-term castration in the male rat. Endokrinologie 76, 23-28. Martinez-Conde E., Leret M. L. and Diaz S. (1985) The influence of testosterone in the brain of the male rat on levels of serotonin (5-HT) and hydroxyindoleacetic acid (5-HIAA). Comp. Biochem. Physiol. 8OC, 41 l-414. McEwen B. S., Pfaff D. W. and Zigmon R. E. (1970) Factors influencing sex hormone uptake by rat brain regions-III. Effects of competing steroid on testosterone uptake. Brain Res. 21, 29-38. Ojeda S. R. and McCann S. M. (1973) Evidence for participation of a catecholaminergic mechanism in the post-castration rise in plasma gonadotropins. NeuroendocrinoloEv 12. 295-3 15. de Paolo-L. V:, McCann S. M. and Negro-Vilar A. (1982) A sex difference in the activation of hypothalamic catecholaminergic and LHRH peptidergic neurons after acute castration. Endocrinology 110, 531-539. Ramirez V. D. (1983) Hormones and striatal dopaminergic activity: a novel neuroendocrine model. In The Anterior Pituitary Gland (Edited by Ajay S. and Bhatnagar), pp. 97-105. Raven Press, New York. Shen J. T. and Ganong W. F. (1976) Effect of Variations in adrenocortical function on dopamine b-hydroxylase and norepinephrine in the brain of the rat. J. Pharmac. exp. Ther. 199, 639448. Simpkins J. W., Kalra P. S. and Kalra S. P. (1980) Effects of testosterone on cathecholamine turnover and LHRH content in the basal hypthalamus and preoptic area. Neuroendocrinology 30, 94100. Tibolt R. E. and Childs G. V. (1985) Cytochemical and cytophysiological studies of gonadotropin releasing hormone (GnRH) target cells in the male rat pituitary: differential effects of androgens and corticosterone on GnRH binding and gonadotropin release. Endocrinology 117, 396-404. Vermes I., Varszegi M., Totti E. K. and Telegdy G. (1979) Action of androgenic steroids on brain neurotransmitters in rats. Neuroendocrinology 28, 386393. Versteed D. H. G., Van Zoest I. and De Kloet E. R. (1984) Acute changes in dopamine metabolism in the media basal hypothalamus following adrenalectomy. Experientia 40, 112-114. Welch A. and Welch B. L. (1969) NE, DA, 5-HT and 5-HTAA in mouse brain. Analyt. Biochem. 30, 161-179.