0013-7227/90/1262-1216$02.00/0 Endocrinology Copyright © 1990 by The Endocrine Society

Vol. 126, No. 2 Printed in U.S.A.

Galanin Is a Physiological Regulator of Spontaneous Pulsatile Secretion of Growth Hormone in the Male Rat* DOMINIQUE M. MAITERt, SHING C. HOOI$, JAMES I. KOENIG, AND JOSEPH B. MARTIN Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114

ABSTRACT. To determine whether galanin (GAL), a 29amino acid neuropeptide, plays a role in the physiological regulation of the pulsatile secretion of GH and PRL in the male rat, secretory patterns of both hormones were studied in freely moving animals after GAL passive immunoneutralization. Adult male Sprague-Dawley rats were equipped with iv and intracerebroventricular catheters. After 7 days, 3 (A of a specific GAL antiserum (GAL-AS) or normal rabbit serum (NRS; controls) were infused in the third ventricle of 10 rats, 25 and 1 h before the animals were bled every 15 min for 6 h (1000-1600 h). Plasma GH and PRL concentrations were measured by RIA, and the hormonal secretory patterns were analyzed by the PULSAR program. Control rats, treated with NRS, displayed typical GH secretion, with pulses of high amplitude (167 ± 27 ng/ml) and low frequency (2.4 ± 0.2 pulses/6 h), separated by periods of low trough levels (3.8 ± 0.6 ng/ml). Rats treated with GALAS had altered pulsatile GH secretion. Pulse height was markedly reduced (77 ± 15 ng/ml; P < 0.01 vs. controls), and peak

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INCE the isolation and characterization of galanin (GAL) from porcine intestine (1), several observations have supported a role for this 29-amino acid peptide as a neurotransmitter or a neuromodulator in the central nervous system (CNS). Indeed, GAL is widely distributed throughout.the CNS (2), especially the hypothalamus (3). Immunohistochemical studies have demonstrated high concentrations of GAL-containing cell bodies in the arcuate, paraventricular, and supraoptic nuclei, while dense GAL-containing fibers emerging from these regions are found in the internal and external layers of the median eminence (ME) (3-5). Specific binding sites for

Received August 30,1989. Address all correspondence and requests for reprints to: Dr. D. Maiter, Unite de Diabetologie et Nutrition, UCL 54.74, Avenue Hippocrate 54, B-1200 Brussels, Belgium. * Presented in part at the 71st Annual Meeting of The Endocrine Society, Seattle, WA, June 21-24, 1989 (Abstract 35). This work was supported by NIH Grants DK-26252 and DK-39251. t Research Assistant of the National Foundation for Scientific Research (Brussels, Belgium) and Recipient of an International Fogarty Award of the NIH (TW-03984). Current address: Unite de Diabetologie et Nutrition, University of Louvain, B-1200 Brussels, Belgium. | Recipient of a Scholarship from the National University of Singapore.

frequency was higher (3.6 ± 0.5 pulses/6 h; P < 0.05), while GH baseline levels and integrated GH secretion over the 6-h sampling period remained unaltered. Injection of rat GH-releasing hormone (1 tig/rat, iv) caused a similar GH stimulation in both groups of rats, as determined by the peak GH response at 5 min (368 ± 112 us. 342 ± 81 ng/ml) or by the integrated GH response over 1 h (5.13 ± 1.30 us. 4.77 ± 1.15 /ig-min/ml in NRS- and GAL-AS-treated rats, respectively; P < 0.05). In contrast to GH, pulsatile secretion of PRL was not affected by the GAL-AS treatment. These results indicate that GAL is a physiological regulator of spontaneous pulsatile secretion of GH, but not PRL, in the male rat. The influence of GAL on GH secretion appears to be exerted within the hypothalamus, mainly by a stimulation of GRF secretion. However, the changes in GH pulse frequency observed after GAL immunoneutralization suggest that GAL might also influence the somatostatin inhibitory tone. (Endocrinology 126: 1216-1222,1990)

GAL have also been identified in the rat hypothalamus and ME (6). In addition, several studies have reported effects of GAL on the hypothalamic-pituitary axis. Pharmacological administration of porcine GAL stimulates the secretion of GH and PRL in rats and humans (7-11) and might reduce the secretion of TSH in rats (12), although this latter effect is controversial (8,13). Intracerebroventricular (icv) injection of the peptide also increases the secretion of LH in ovariectomized rats, but only when they are primed with estrogen and progesterone (14). Recent data from our laboratory also indicate that GAL might be an important component in the regulation of ACTH (13) and arginine vasopressin secretion in the rat (15). Despite this increasing number of studies, a physiological role for GAL in the brain has not yet been established. Preliminary data have been recently reported, showing that GAL passive immunization in the rat affects steady state levels of GH, PRL, and TSH (12). However, effects of GAL antiserum (GAL-AS) on the pulsatile patterns of GH and PRL secretions were not addressed in that study.

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GAL REGULATES PULSATILE GH SECRETION The present work was intended to demonstrate a physiological role for GAL in the regulation of pulsatile GH and PRL secretion by studying the effects of icv administration of GAL-AS on both hormonal secretions in unstressed, freely moving male rats. The GH response to rat GH-releasing hormone (rGRF) was also assessed in order to establish the site of action of the GAL-AS.

Materials and Methods Animals and experimental procedures Adult male rats (200-250 g) of the Sprague-Dawley strain (Charles River, Wilmington, MA) were maintained in a temperature (22 ± 1 C)- and light-controlled (lights on from 07001900 h) animal facility. Standard rat chow pellets and tap water were provided ad libitum. Under pentobarbital anesthesia (50 mg/kg, ip), a Silastic cannula was placed into the right atrium via the external jugular vein, as previously described (16), and a permanent stainless steel catheter (23 gauge) was implanted stereotaxically into the third cerebral ventricle (icv cannula) using coordinates reported by Paxinos and Watson (17). After surgery, the animals were housed in individual cages and handled daily to minimize stress. Jugular cannulae were kept patent by flushing with 0.2 ml heparinized saline (50 IU/ml) every other day. Studies were performed only on rats that had regained preoperative body weight. One week after surgery, 3 fi\ rabbit GAL-AS or normal rabbit serum (NRS; controls) were microinfused through the icv cannula at 0900 h, on 2 consecutive days. Each experimental group consisted of 10 rats. One hour after the second icv injection, blood samples (0.6 ml) were collected every 15 min from 1000-1600 h through the iv cannula. Plasma was immediately separated by centrifugation and stored at -20 C until assayed for rGH and rPRL. The remaining blood cells were resuspended in 0.3 ml heparinized saline (30 IU/ml) and reinfused after the next blood sampling. At the end of the sampling period (1600 h), 1 fig rGRF (Peninsula Laboratories, Belmont, CA) was injected iv, and blood samples (0.4 ml) were withdrawn 5, 15, 30, and 60 min after injection. Rats were then killed, and the position of the icv cannula was verified in each animal by dye injection. Antiserum The GAL-AS was generated in a rabbit using synthetic porcine GAL (Peninsula Laboratories), coupled to BSA with carbodiimide and has been previously characterized (18). The antiserum does not cross-react with other peptides, including GRF, somatostatin, neuropeptide-Y, vasoactive intestinal polypeptide, secretin, leucine-enkephalin, substance-P, and LHRH. Also, displacement curves of [125I]GAL binding by rat brain extracts and the synthetic porcine GAL standard curve exhibit parallelism (18). RIAs Plasma concentrations of rGH and rPRL were determined by RIAs using materials provided by the National Hormone and Pituitary Program. Results were expressed in terms of rGH

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RP-2 and rPRL RP-3 standards, respectively. Sensitivity and intra- and interassay coefficients of variation for the rGH assay were 1.0 ng/ml, 9.8%, and 11%, and those for the rPRL assay were 0.5 ng/ml, 9.6%, and 14.1%, respectively. Values below the sensitivity of the assay were treated as the minimum detectable level for statistical analysis. Values above the range of the assay were reassayed with appropriate dilution. Statistical analysis The pulsatile patterns of GH and PRL secretion were analyzed in individual animals using the PULSAR program (19) and a parameter setting similar to the one described by Jansson and Frohman (20) in order to fit the well known characteristics of the GH secretory pattern in normal male rats (16). Exclusion criteria for pulse identification were 3.8, 2.6, 1.9, 1.5, and 1.2 within-assay SD for pulses with a duration of 1, 2, 3, 4, and 5 time points, respectively. The within-assay SD was determined by assaying in quintuplet six samples with hormone concentrations distributed across the assay range. The linear regression equations relating the SD (y) to hormonal concentration (JC) were, for rGH: y = 0.26 + 0.08x, and for rPRL: y = -0.01 + O.lOx. A peak-splitting criterion of 6 SD allowed the program to analyze the large multiplasic GH surges as single secretory phenomena, rather than separate peaks. The amount of hormone released during the 6-h sampling period or during the hour after the rGRF injection were determined by calculation of the area under the curves. Values are expressed as the mean ± SE. Differences between groups were analyzed by the nonparametric Mann-Whitney U test, with P < 0.05 considered statistically significant. Results Control (NRS-treated) male rats displayed a typical rhythm of GH secretion (Fig. 1). High amplitude GH pulses (81-515 ng/ml) occurred regularly, with a mean interpulse interval (interval between the peaks) of 166 ± 10 min. Between the pulses, basal GH secretion was low, with levels often lower than 3 ng/ml. Intracerebroventricular administration of a specific GAL-AS, 25 and 1 h before the start of the sampling period, significantly altered the GH secretory pattern in male rats (Fig. 1). The GH pulses were of lower amplitude (21-194 ng/ml) and shorter duration in the GAL-AS-injected rats than in the control animals (74 ± 8 vs. 95 ± 5 min, respectively; P < 0.05). Furthermore, pulses occurred more frequently and less regularly after GAL passive immunization, with a mean interpulse interval of 111 ± 21 min (P < 0.01 us. controls). PULSAR analysis of the GH secretory patterns confirmed that the main alterations caused by the icv infusion of GAL-AS were a 55% decrease in GH pulse height (increment over baseline) and a 50% increase in GH pulse frequency, without any significant effect on GH baseline levels (Fig. 2). Integrated GH secretion over the 6-h sampling period was decreased by 27% in GALAS-treated rats, but this reduction did not reach statis-

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Endo • 1990 Voll26«No2

GAL REGULATES PULSATILE GH SECRETION

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GALANIN ANTI SERUM

CONTROLS (213)

(187)

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GH PULSE FREQUENCY (no./6hr) *

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*

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BASELINE GH (ng/ml)

INTEGRATED GH SECRETION 20 i(Ug . mtn/ml)

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CLOCK TIME (hours) FIG. 1. Representative plasma GH profiles in individual male rats, 25 and 1 h after icv injection of 3 fi\ NRS (controls; left panel) or a specific antiserum against porcine GAL (right panel). GH peaks identified by the PULSAR program are indicated by single arrows (one time point peaks) or connected arrows (multiple time points peaks). Plasma GH values greater than 150 ng/ml (RP-2 standard) are shown in parenthe-

tical significance. GAL-AS effects on GH secretion were still present at the end of the sampling period. A similar 55% decrease in GH pulse height occurred after GAL immunoneutralization whether we considered the first pulse (139 ± 42 vs. 61 ± 11 ng/ml; P < 0.05) or the last pulse detected by the PULSAR analysis (151 ± 30 vs. 71 ± 18 ng/ml, controls and GAL-AS-treated rats, respectively; P < 0.05). Intravenous injection of rGRF (1 /ig/rat) caused a similar stimulation of GH release in controls and rats injected with the GAL-AS (Fig. 3). Peak GH responses to GRF were 368 ±112 ng/ml and 342 ± 81 ng/ml, and integrated GH responses over 1 h were 5.13 ± 1.30 and 4.77 ± 1.15 /ug-min/ml in NRS- and GAL-AS-treated rats, respectively (P > 0.05). In both controls and GAL-AS-injected male rats, the PRL secretory pattern was characterized by the emergence of two or three small pulses (2.4-26.0 ng/ml), usually in the afternoon period (Fig. 4). There was no significant effect of GAL immunoneutralization on mean

19

10

FIG. 2. Characteristics of the GH secretory pattern in male rats injected with 3 n\ NRS (controls; n = 10). Data were obtained from the PULSAR analysis of plasma GH profiles (pulse height and frequency; baseline GH) or calculation of the area under the GH curve (integrated GH secretion). Values are shown as the mean ± SE. *, P < 0.05; **, P < 0.01 (vs. NRS-treated controls).

PRL pulse height, pulse frequency, basal secretion, or integrated PRL secretion over the 6-h sampling period (Fig. 5). Discussion Previous studies have shown that pharmacological administration of porcine GAL into the CNS, in doses ranging from 15-600 pmol (50-2000 ng) produces a rapid and dose-dependent increase in plasma GH levels in male (8, 9, 12) and ovariectomized female rats (7). Also, peripheral iv injection of the neuropeptide at higher doses

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GAL REGULATES PULSATILE GH SECRETION

• O

0

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• CONTROLS (NRS) O GALANIN AKTISERUM

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GALANIN ANTISERUM

PRL PULSE HEIGHT (ng/ml) 10

PRL PULSE FREQUENCY (no./6hr) 3r

r

30

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FiG. 3. Plasma GH response to a bolus iv injection of rGRF (1 Mg/rat) in NRS-treated control male rats ( • - • ) and rats previously injected icv with 3 n\ GAL-AS (O- -O). BASELINE PRL (ng/ml)

3r

GALANIN ANTISERUM

CONTROLS

•

INTEGRATED PRL SECRETION 1.3 r(ug • mln/ml)

n 1.0

i

o.s

FIG. 5. Characteristics of the PRL secretory pattern in male rats injected with 3 ^1 NRS (controls; n = 10) or a specific GAL-AS (n = 10). Data were obtained from the PULSAR analysis of plasma PRL profiles (pulse height and frequency; baseline PRL) or calculation of the area under the PRL curve (integrated PRL secretion). Values are shown as the mean ± SE. No statistical difference was observed between the two experimental groups of rats.

CLOCK TIME (hours) FiG. 4. Representative plasma PRL profiles in individual male rats, 25 and 1 h after icv injection of 3 IJ.1 NRS (controls; left panel) or a specific GAL-AS (right panel). Symbols are described in Fig. 1.

stimulates GH release in rats (12) and humans (11, 21). In the present study we demonstrate that immunoneutralization of endogenous GAL within the CNS significantly disrupts the normal GH secretory pattern in male rats, without affecting the pituitary GH response to exogenous GRF administration. These results support an important physiological role for endogenous GAL in the control of spontaneous pulsatile GH secretion in the male rat. They also extend the results recently reported by Ottlecz et al. (12), showing a prolonged inhibition of mean steady state GH secretion after icv injection of a different GAL-AS.

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GAL REGULATES PULSATILE GH SECRETION

The major impairment in GH secretion induced by GAL-AS injection is related to a severe reduction in GH pulse amplitude, a parameter which appears to be critical for optimal growth-promoting actions of GH, such as the liver production of insulin-like growth factor-I or the stimulation of body growth (22). Other alterations of the GH pulses include an increase in pulse frequency and a loss of the normal 3-h periodicity. We confirm that the net effect of GAL passive neutralization is inhibitory on integrated GH secretion, but this inhibition was only partial (27%), probably because GH secretion is still maintained by the activity of other neurotransmitters known to regulate the GH axis (23). However, we cannot rule out the possibility that neutralization of endogenous rat GAL within the brain has been incomplete, since our antiserum was raised against the porcine peptide (18), and the sequences of the two heterologous peptides differ by three amino acids near the C-terminal end (24). Nevertheless, the antibody is directed against the midportion of the molecule and is capable of reacting with rat GAL, since the displacement of GAL tracer by rat brain extracts and the synthetic porcine peptide are parallel (18). Moreover, we have demonstrated previously that our antiserum is effective in detecting brain GAL immunoreactivity by both RIA and immunocytochemistry (25). Furthermore, as in other immunoneutralization studies, the incomplete suppression of GH secretion could be due to the required diffusion of the GAL-AS into the brain before its action can be exerted. More frequent or continuous administration of GAL-AS was avoided in the study, because of the possibility of stressing the animals, which could interfer with the normal episodic secretion of GH and PRL. Also, long-lasting effects of the antiserum are supported by the observation of a similar decrease in GH pulse height at the beginning and end of the sampling period in the rats infused with the GAL-AS. Although the precise mechanisms involved in GAL regulation of GH secretion remain to be determined, current evidence suggests that the neuropeptide is primarily acting in the hypothalamus and/or ME. High concentrations of GAL and GAL-binding sites are found in these areas (2-6), and the GH-releasing effects of GAL are much more pronounced when administered icv rather than iv (12). In addition, GAL has little or no effect on basal or stimulated GH release from anterior pituitary cells in vitro (7, 12, 26). The small volume of GAL-AS infused icv in the present study and its inability to alter the GH response to GRF stimulation also point toward a hypothalamic site of action and argue against the possibility that the centrally administered antiserum could have exerted its effects at the pituitary level, after diffusion into the hypophyseal circulation. It remains to be demonstrated whether pituitary GAL also regulates

Endo • 1990 Voll26«No2

the secretion of GH, especially in the presence of high circulating levels of estrogens, which strongly stimulate the synthesis and secretion of the neuropeptide in the adenohypophysis (27). Episodic GH secretion in male rats is governed by the interactions between two hypothalamic hormones (28, 29). GRF stimulates GH secretion and mainly determines the high amplitude of the GH pulses, while somatostatin exerts an inhibitory influence, which is responsible for the low interpulse secretion and the GH pulse frequency, since a GH pulse occurs only if the somatostatin tone is withdrawn (28). Most of the neurotransmitters modulating GH secretion act via one of these two hypothalamic factors (23, 29). The data reported here suggest that GAL might influence the secretion of both GH-regulating hormones. On the one hand, GAL probably has a stimulatory action on hypothalamic and/or ME GRF activity, as we observed a severe decrease in GH pulse amplitude together with a normal GH response to GRF injection after central GAL immunoneutralization. Indeed, alterations at the pituitary level or an increase in the somatostatin inhibitory action would reduce the GH pulse height, but also decrease somatotroph responsiveness to GRF. An effect of GAL on GRF secretion has already been proposed by Murakami et al. (30), who have observed that treatment of male rats with specific GRF antiserum markedly inhibits the GH response to either icv or iv GAL administration. Concomitantly, they showed that GAL might also act through a stimulation of central a-adrenergic and yaminobutyric acid-ergic systems, which, in turn, activate GRF pathways and GH secretion (30). A similar mechanism, involving presynaptic activation of catecholaminergic neurons and stimulation of GRF activity, also seems to account, at least in part, for the GH-releasing effect of GAL in infant rats (31). However, the coexistence of GAL and GRF in the same arcuate neurons has been demonstrated, thereby enhancing the likelihood of direct interactions between these two peptides (32). In humans, the GH response to exogenous GRF infusion is markedly enhanced by GAL, indicating that one possible mode of action for GAL would be an inhibition of hypothalamic somatostatin release (21). Such a mechanism does not seem to be predominant in the rat, since neither basal GH secretion nor the GH response to GRF injection is affected by GAL-AS injection. However, this treatment increases the GH pulse frequency, and this effect could be explained by a more subtle influence of GAL on the periodicity of the somatostatin inhibitory tone within the hypothalamic-pituitary axis. Another possible mechanism would be a direct effect on a pulse generation system, for example at the level of the suprachiasmatic nucleus, where GAL cell bodies and fibers are found in appreciable amounts (4). This might well ex-

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GAL REGULATES PULSATILE GH SECRETION plain the loss of the regular 3-h periodicity of the GH pulses after GAL neutralization. Our data do not support an important contribution of GAL in the physiological control of pulsatile PRL release in the male rat, as GAL-AS administration was without any demonstratable effect on this hormonal secretion. Accordingly, GAL stimulates PRL release in rats, only when large doses are used and administered directly into the CNS (10,12). However, it remains possible that GAL mediates the PRL response to some stimulatory or inhibitory signals. Indeed, it has been shown that GAL is able to stimulate the release of vasoactive intestinal polypeptide, a PRL-releasing factor, from hypothalamic fragments in vitro (33) and to inhibit the release of dopamine, a PRL-inhibiting factor, from ME nerve terminals (34). In conclusion, hypothalamic GAL appears to be a determining factor in the control of spontaneous pulsatile secretion of GH in the male rat. This neuropeptide regulates both the high amplitude of the GH pulses, most likely through a stimulation of GRF secretion, and the regular 3-h cycle frequency of the GH rhythm, perhaps by influencing the periodicity of the somatostatin inhibitory tone. Therefore, it is tempting to speculate that GAL might be an important factor involved in the development and/or the maintenance of the normal GH secretory pattern in male rats and, hence, in the optimal growth of the animal. A modulation by GAL of the numerous metabolic and immune functions that are under GH control (35, 36) could also be hypothesized, although there is no evidence yet that normal GH pulsatility is required for the regulation of these pathways byGH.

Acknowledgments We wish to thank Ms. Judith Audet-Arnold and Carol Milbury for expert technical assistance, Ms Denise Heavern for helpful assistance in the PULSAR analysis of hormonal secretory patterns, and Mrs. Elisabeth Thompson, Ms. Sharon Melanson, and Ms. Nicole AmatPeiro for their excellent secretarial work. We are also grateful to Dr. S. Raiti and the National Hormone and Pituitary Program for providing the materials used in the rat GH and PRL RIAs.

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galanin binding sites in the rat central nervous system. Eur J Pharmacol 124:381 7. Ottlecz A, Samson WK, McCann SM 1986 Galanin: evidence for a hypothalamic site of action to release growth hormone. Peptides 7:51 8. Melander T, Fuxe K, Harfstrand A, Eneroth P, Hokfelt T 1987 Effects of intraventricular injections of galanin on neuroendocrine functions in the male rat. Possible involvement of hypothalamic catecholamine neuronal systems. Acta Physiol Scand 131:25 9. Murakami Y, Kato Y, Koshiyama H, Inoue T, Yanaihara N, Imura H 1987 Galanin stimulates growth hormone (GH) secretion via GH-releasing factor (GRF) in conscious rats. Eur J Pharmacol 136:415 10. Koshiyama H, Kato Y, Inoue T, Murakami Y, Ishikawa Y, Yanaihara N, Imura H 1987 Central galanin stimulates pituitary prolactin secretion an rats: possible involvement of hypothalamic vasoactive intestinal polypeptide. Neurosci Lett 75:49 11. Bauer FE, Ginsberg L, Venetikou M, McKay DJ, Burrin JM, Bloom SR 1986 Growth hormone release in man induced by galanin, a new hypothalamic peptide. Lancet 2:192 12. Ottlecz A, Snyder GD, McCann SM 1988 Regulatory role of galanin in control of hypothalamic-anterior pituitary function. Proc Natl Acad Sci USA 85:9861 13. Hooi SC, Maiter D, Koenig JI, Martin JB, Effects of intracerebroventricular injection of galanin antiserum (GAL-AS) on plasma adrenocorticotrophic (ACTH) and thyroid-stimulating (TSH) hormones in the rat. 71st Annual Meeting of The Endocrine Society, Seattle WA, 1989, p 398 (Abstract) 14. Sahu A, Crowley WR, Tatemoto K, Balasubramaniam A, Kalra SP 1987 Effects of neuropeptide Y, NPY analog (norleucine 4NPY), galanin and neuropeptide K on LH release in ovariectomized (OVX) and OVX estrogen, progesterone-treated rats. Peptides 8:921 15. Koenig JI, Hooi S, Gabriel SM, Martin JB 1989 Potential involvement of galanin in the regulation of fluid homeostasis in the rat. Regul Peptides 24:81 16. Tannenbaum GS, Martin JB 1976 Evidence for an endogenous ultradian rhythm governing growth hormone secretion in the rat. Endocrinology 98:562 17. Paxinos G, Watson C 1986 The Rat Brain in Sterotoxic Coordinates, ed 2. Academic Press, Orlando, p 21 18. Gabriel SM, McGarvey UM, Koenig JI, Swartz KJ, Martin JB, Beal MF 1988 Characterization of galanin-like immunoreactivity in the rat brain: effects of neonatal glutamate treatment. Neurosci Lett 87:114 19. Merriam GR, Wachter KW 1982 Algorithms for the study of episodic hormone secretion. Am J Physiol 243:E310 20. Jansson JO, Frohman LA 1987 Differential effects of neonatal and adult androgen exposure on the growth hormone secretory pattern in male rats. Endocrinology 120:1551 21. Davis TME, Burrin JM, Bloom SR 1987 Growth hormone (GH) release in response to GH-releasing hormone in man is 3-fold enhanced by galanin. J Clin Endocrinol Metab 65:1248 22. Maiter D, Underwood LE, Maes M, Davenport ML, Ketelslegers JM 1988 Different effects of intermittent and continuous growth hormone (GH) administration on serum somatomedin-C/insulinlike growth factor I and liver GH receptors in hypophysectomized rats. Endocrinology 123:1053 23. Martin JB, Reichlin S 1987 Clinical Neuroendocrinology, ed 2. Davis, Philadelphia, p 243 24. Kaplan LM, Spindel ER, Isselbacher KJ, Chin WW 1988 Tissuespecific expression of the rat galanin gene. Proc Natl Acad Sci USA 85:1065 25. Gabriel SM, Marshall PE, Martin JB 1988 Interactions between growth hormone releasing hormone, somatostatin and galanin in the control of growth hormone secretion. In: Bercu BB (ed) Basic and Clinical Aspects of Growth Hormone. Plenum Press, New York, p 73 26. Gabriel SM, Milbury CM, Nathanson JA, Martin JB 1988 Galanin stimulates rat pituitary growth hormone secretion in vitro. Life Sci 42:1981 27. Kaplan LM, Gabriel SM, Koenig JI, Sunday ME, Spindel ER,

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Elde R, Goldstein M, Hemmings H, Ouimet C, Walaas I, Greengard P, Vale W, Weber E, Wu Y-J, Chang K-J 1986 The hypothalamic arcuate nucleus-medien eminence complex: immunohistochemistry of transmitters, peptides and DARPP-32 with special reference to coexistence in dopamine neurons. Brain Res Rev 11:97 Inoue T, Kato Y, Koshiyama H, Yanaihara N, Imura H 1988 Galanin stimulates the release of vasoactive intestinal polypeptide from perfused hypothalamic fragments in vitro and from periventricular structures into the cerebrospinal fluid in vivo in the rat. Neurosci Lett 85:95 Nordstrom O, Melander T, Hokfelt T, Bartfai T, Goldstein M 1987 Evidence for an inhibitory effect of the peptide galanin on dopamine release from the rat median eminence. Neurosci Lett 73:21 Davidson MB 1987 Effect of GH on carbohydrate and lipid metabolism. Endocr Rev 8:115 Berczi I 1986 Immunoregulation by pituitary hormones. In: Berczi I (ed) Pituitary Function and Immunity. CRC Press, Boca Raton, p227

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