| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
ARTICLES |
Institute of Experimental and Clinical Pharmacology and Toxicology, Medical University Lübeck, D-23538 Lübeck, Germany
Address all correspondence and requests for reprints to: Olaf Jöhren, Ph.D., Institute of Experimental and Clinical Pharmacology and Toxicology, Medical University Lübeck, Ratzeburger Allee 160, D-23538 Lübeck, Germany. E-mail: joehren{at}medinf.mu-luebeck.de
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Orexin (hypocretin)-immunoreactive fibers project to various brain regions, such as the cerebral cortex, limbic system, locus coeruleus, and brainstem, and to the spinal chord (4, 5, 6, 7). The orexin receptor subtypes OX1 and OX2 are also widely expressed in the rat brain, and the expression patterns are in good agreement with the distribution of orexin-immunoreactive fibers (8). Based on the widespread projections of orexinergic neurons and functional considerations on the innervated brain areas, the involvement of orexins in the control of functions other than feeding behavior was proposed, including the control of neuroendocrine systems and the autonomic nervous system. Subsequently, orexins were shown to increase plasma ACTH and corticosterone levels and to reduce plasma PRL and GH levels when injected intracerebroventricularly (icv) (9, 10, 11). Furthermore, icv injected orexin increased or reduced plasma LH levels in female rats depending on the status of ovarian steroids (12, 13). In addition, icv or intracisternally injected orexin increased mean arterial blood pressure, heart rate, renal sympathetic nerve activity, and plasma catecholamine levels (14, 15, 16). Moreover, in dogs, mice, and humans the orexin system seems to be closely associated with the pathogenesis of narcolepsy (17, 18, 19).
To date, orexins were considered to be exclusively synthesized in the brain (20). However, recent results indicated endocrine effects after peripheral administration of orexins (21, 22, 23). On the other hand, iv injected orexin A is able to cross the blood-brain barrier (24), and the sites of peripheral orexin generation and the distribution of orexin receptor subtypes are not yet established. Therefore, we analyzed the expression of prepro-orexin as well as OX1 and OX2 receptor mRNA in various tissues of adult male and female rats.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
For in situ hybridization and immunohistochemistry, tissues were obtained from an extra set of animals (n = 3), and sections (16 µm) were cut at -20 C in a cryostat. The sections were mounted on aminoalkylsilane-coated glass slides (Sigma, St. Louis, MO) and stored at -80 C. Every 10th section was stained with hematoxylin/eosin.
RNA isolation and cDNA synthesis
Tissue samples were homogenized in the presence of guanidinium
isothiocyanate, and total RNA was isolated using silica gel-based spin
columns (RNeasy Kit, QIAGEN, Hilden, Germany). Genomic DNA
was digested by thorough treatment with deoxyribonuclease I
(QIAGEN). First strand cDNA was synthesized from 1 µg
total RNA in a volume of 20 µl containing 5 mM
MgCl2, 10 mM Tris-HCl (pH 9.0), 50
mM KCl, 0.1% Triton X-100, 1 mM deoxy-NTPs,
1U/µl RNasin, 0.5 µg oligo(deoxythymidine)15
primer, and 15 U AMV reverse transcriptase (Promega Corp., Mannheim, Germany). For no-template control samples, RNA
was replaced by H2O. Possible contamination of
RNA samples with genomic DNA was monitored by omission of the reverse
transcriptase during the synthesis of the first strand cDNA. No genomic
DNA was detected under these conditions.
PCR and cDNA cloning
Prepro-orexin, OX1 and
OX2 receptors, and ß-actin-specific sense and
antisense oligonucleotide primers were designed based on the published
cDNA sequences (1, 26) using the Primer3 software
developed by S. Rozen and H. J. Skaletsky (code available at
http://www-genome.wi.mit.edu/genome_software/other/primer3.html).
Oligonucleotides were obtained from Live Technologies (Karlsruhe,
Germany). The oligonucleotide sequence and product size for each primer
pair used are shown in Table 1
.
|
The amplified prepro-orexin and OX1 and OX2 receptor cDNA fragments were subcloned into the pCR-II vector using a TA cloning kit (Invitrogen, Groningen, The Netherlands). The identity of the cloned cDNAs was confirmed by nucleotide sequencing (MWG Biotech, Ebersberg, Germany).
Real-time quantitative RT-PCR
Real-time quantitative RT-PCR was performed on the GeneAmp 5700
sequence detection system (PE Applied Biosystems,
Weiterstadt, Germany) using a SYBR green I reaction system (Eurogentec
SA, Seraing, Belgium). Sense and antisense primers were identical to
those described above (Table 1
). Each sample was analyzed in triplicate
along with standards and no-template controls. The reaction contained
100 ng cDNA, 0.3 µM primers, 10 mM Tris-HCl,
50 mM KCl, 3 mM MgCl2,
0.2 mM deoxy-NTPs, and 1.25 U Hot GoldStar DNA Polymerase
(Eurogentec, Cologne, Germany) in a final volume of 50 µl. After
denaturation at 95 C for 10 min, the cDNA products were amplified for
40 cycles, each cycle consisting of denaturation at 95 C for 15 sec,
annealing at 56 C for 30 sec, and extension at 60 C for 30 sec. Product
purity was confirmed by dissociation curve and agarose gel
analysis.
Copy number calculations were based on the cycle threshold method
(27). Serial dilutions of known amounts of the cloned
prepro-orexin and OX1 and
OX2 receptor cDNA fragments were used to generate
standard curves. The threshold cycle number for each sample was
calculated using the GeneAmp 5700 sequence detection system software
with an automatic baseline setting and a fluorescence threshold
(Rn) of 0.3. No-template control samples
typically did not surpass the Rn of 0.3 at 40
cycles (Fig. 3, A and E). Only in the case of OX1
receptor mRNA amplification did no-template controls reach the
Rn value of 0.3 at 35 about cycles (Fig. 3C).
Therefore, copy numbers of mRNA were generally considered
nonsignificant if the threshold cycle number exceeded 35 cycles. The
value corresponds to a detection limit of about 20 mRNA copies/100 ng
cDNA. For comparison of male and female rats, copy numbers of
OX1 and OX2 receptor mRNA
were normalized to ß-actin mRNA levels.
|
Sections were fixed for 10 min in 4% paraformaldehyde in PBS (0.5 M NaCl and 0.1 M phosphate buffer, pH 7.4) followed by two washes in PBS. The sections were then dehydrated in 70% (2 min) and 95% (2 min) ethanol and air-dried. Sections were then hybridized for 16–18 h at 42 C with 1–2 pmol/ml labeled oligonucleotides in hybridization buffer containing 50% formamide, 10% dextran sulfate, 1 x Denhardt’s solution, 20 mM Tris (pH 7.5), 0.3 M NaCl, 1 mM EDTA, 150 mM dithiothreitol, 0.2% SDS, 100 µg/ml salmon sperm DNA, and 250 µg/ml yeast tRNA. After hybridization, sections were washed several times in 1 x NaCl-sodium citrate (SSC) at room temperature, followed by high stringency washing in 1 x SSC at 55 C for 30 min. After washing in 1 x SSC and 0.1 x SSC for 1 min each at room temperature, sections were dehydrated through a series of ethanols containing 300 mM ammonium acetate, air-dried, and exposed to Hyperfilm 3H (Amersham Pharmacia Biotech) for 1–2 wk. The specificity of the hybridization signal was assessed by hybridization of the 33P-labeled antisense oligonucleotides in the presence of a 100-fold molar excess of the respective unlabeled oligonucleotide.
Immunohistochemistry
To distinguish the area of adrenal OX2
receptor expression obtained by in situ hybridization, we
immunohistochemically stained the adrenal medulla adjacent sections
using a monoclonal antibody against tyrosine hydroxylase (clone TH-2,
Sigma, Deisenhofen, Germany). This antibody was shown
previously to specifically stain catecholaminergic neurons
(28). After fixation of the sections in ice-cold acetone
for 10 min, the sections were incubated with the primary antibody
(1:2000 dilution) for 60 min at 37°C in Tris-buffered saline, pH 7.6,
containing 0.1% BSA. Positive staining was detected by the
avidin-biotin complex method (29) with a biotinylated
secondary antibody and peroxidase-conjugated streptavidin, using the
LSAB kit (DAKO Corp., Hamburg, Germany) according to the
manufacturer’s protocol. The chromogen was 3,3-diaminobenzidine
tetrahydrochloride (Sigma). As a negative control,
sections were processed for immunostaining in the absence of the first
antibody. In these control experiments no staining was observed.
RIA
Orexin A was extracted from plasma samples by adsorption to
phenyl-silica (Isolute SPE, International Sorbent Technology, Mid
Glamorgan, UK). After elution in 60% acetonitrile and 1%
trifluoroacetic acid, the samples were concentrated by lyophilization.
The recovery of [125I]orexin A by this
procedure was more than 90%. Orexin A concentrations were determined
in the reconstituted samples by a specific RIA according to the
instructions of the manufacturer (Peninsula Laboratories, Inc., Belmont, CA). The detection limit of the assay, based on
the amount of orexin A that produced at least 10% tracer displacement,
was 3 pg/tube. All samples were analyzed within the same assay for
which an intraassay variability of 9% was calculated.
Statistical analysis
Data are presented as the mean ± SEM.
Differences between male and female rats were evaluated by t
test. P < 0.05 was considered statistically
significant.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
). Brain samples were used as
positive controls, because mRNAs for prepro-orexin as well as those for
the orexin receptor subtypes OX1 and
OX2 are present in rat brain (1, 3, 8).
|
). In one case, we
detected very low levels of prepro-orexin mRNA in heart. No
prepro-orexin mRNA was detected in pituitary, adrenal, kidney, stomach,
duodenum, jejunum, pancreas, liver, spleen, aorta, lung, thyroid,
adipose tissue, and muscle of male rats (Fig. 1B), and no prepro-orexin
was detected in ovaries (Fig. 2). Using
real-time quantitative PCR we confirmed very high mRNA levels of
prepro-orexin in the rat brain (300,000 copies/100 ng cDNA) and the
presence of prepro-orexin mRNA in testes (Fig. 3, A and B). In ovary and heart,
amplification above the threshold value of 0.3 occurred only at cycle
numbers higher than 35 and was considered nonsignificant.
|
). OX1
receptor mRNA levels were highest in the brain, followed by lower
levels in the pituitary and very low levels in the kidney, thyroid
gland, testes, ovaries, and adrenals (Fig. 3, C and D).
OX2 receptor-specific DNA fragments (314 bp) were
amplified from RNA of the rat brain, lung, adrenal, and pituitary
(Fig. 1D). The highest amounts of OX2
receptor mRNA was found in the adrenal, which was 4 times higher than
brain OX2 receptor mRNA levels (Fig. 3, E and F).
In lung and pituitary, significant, but low, levels of
OX2 receptor mRNA were present (Fig. 3F). No
significant amounts of OX1 and
OX2 receptor mRNA were detected in stomach,
duodenum, pancreas, liver, spleen, heart, aorta, adipose tissue, or
muscle.
The specificity of the RT-PCR was verified by restriction analysis
(Fig. 4). Restriction of the amplified
prepro-orexin fragments with PvuII resulted in the expected
cleavage into two discrete fragments (Fig. 4A) as predicted from the
rat cDNA sequence (1). The specificity of
OX1 and OX2 receptor mRNA
amplification was confirmed by restriction with AluI (Fig. 4, B and C). For a final proof of the RT-PCR specificity, the
nucleotide sequences of subcloned amplification fragments of
prepro-orexin and OX1 and
OX2 receptor cDNA were determined by sequence
analysis and were found to be identical to the corresponding
prepro-orexin and OX1 and
OX2 receptor cDNA sequences (1).
|
). No amplification
product was detected when reverse transcriptase was omitted during cDNA
synthesis (Fig. 1A). Moreover, control reactions without cDNA samples
that were run in parallel in all PCR assays resulted in low background
fluorescence (Fig. 3). Therefore, the presence of genomic DNA in our
samples and DNA contaminations of the reactions from other sources can
be excluded.
Localization of OX2 receptor mRNA in the adrenal gland
by in situ hybridization
To further explore the very high levels of
OX2 receptor mRNA in the rat adrenal, we used
radiolabeled antisense oligonucleotides to localize mRNA expression by
in situ hybridization. As orexins have been shown to
regulate tyrosine hydroxylase mRNA, the rate-limiting enzyme in
catecholamine synthesis, in rat pheochromocytoma cells
(30) we specifically immunostained the adrenal medulla
using an antibody against tyrosine hydroxylase to distinguish between
OX2 receptor mRNA expression in the medulla and
that in the adjacent zona reticularis (Fig. 5B). However, no
OX2 receptor mRNA was detected in the adrenal
medulla (Fig. 5D), where tyrosine hydroxylase-like immunoreactivity is
present in adjacent sections (Fig. 5B). Conversely, high levels of
OX2 receptor mRNA were found in the zona
glomerulosa and reticularis, but not in the zona fasciculata of the
adrenal cortex (Fig. 5D). The specificity of the hybridization signals
was shown by the absence of hybridization signals in the presence of
excessive unlabeled antisense oligonucleotides (Fig. 5, E and F). No
significant levels of OX1 receptor mRNA were
detected in the adrenal gland (Fig. 5C), although a strong signal was
obtained in the locus coeruleus in simultaneously hybridized brain
sections that served as positive controls (not shown). Possibly due to
the low expression levels we could not detect OX1
or OX2 receptor mRNA by in situ
hybridization with antisense oligonucleotide probes in other peripheral
tissues, such as pituitary, kidney, and testes (not shown).
|
). No differences between male
and female rats were found in the amounts of hypothalamic
OX2 receptor mRNA (Fig. 6A). In male rats,
OX1 receptor mRNA levels in the pituitary (Fig. 6B) and OX2 receptor mRNA levels in the adrenal
(Fig. 6C) were more than 5-fold higher compared with those in female
rats. In situ hybridization with radiolabeled
OX2 receptor antisense oligonucleotides confirmed
the comparatively low OX2 receptor expression in
the rat female adrenal gland (Fig. 7).
|
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
The expression of OX1 and OX2 receptors in pituitary and adrenal indicates a possible role of orexins in the regulation of endocrine systems, in particular of the hypothalamic-pituitary-adrenal axis. The effects of icv injected orexins on plasma levels of ACTH and corticosterone have been shown previously (9, 10, 11). Our present results indicate that orexins may also regulate pituitary and adrenal hormones by direct action on these organs. The localization of OX2 receptor mRNA in the zona glomerulosa and reticularis of the adrenal cortex supports the finding by Malendowitz et al. (22) of increased plasma corticosterone levels after sc injection of orexins and enhanced corticosterone secretion from dispersed rat adrenocortical cells by orexins. Recent results also demonstrated the stimulatory effect of orexin on the release of cortisol from human adrenal glands (33). As we found high levels of OX2 receptor mRNA, but could not detect OX1 receptor mRNA in the rat adrenal gland by in situ hybridization and only very low OX1 receptor mRNA levels by the highly sensitive real-time PCR, the effects of orexins on corticosterone synthesis and release in rats might be mediated mainly via the OX2 receptor.
Orexins have been shown to decrease tyrosine hydroxylase mRNA levels and dopamine release from the PC12 rat pheochromocytoma cell line (30), and OX1 as well as OX2 receptor-like immunoreactivity has been found in the adrenal medulla of rats (31). In contrast, we could not detect orexin receptor mRNA in the adrenal medulla by in situ hybridization. However, there may be low levels of orexin receptor mRNA in the adrenal medulla, and beside the stimulation of corticosterone release orexins may influence adrenal catecholamine synthesis and release.
The described effects of orexins on plasma levels of pituitary hormones such as ACTH, GH, PRL, and LH occurred after the injection of orexins into the brain ventricles, and these effects are most likely mediated via the corresponding releasing hormones (9, 11, 12, 13). Plasma TSH levels decreased after iv injection of orexin (34). However, this effect may not be a direct effect on the pituitary, but appears to arise from inhibition of TRH secretion in the hypothalamus (34). Thus, the physiological roles of orexin receptors in tissues such as pituitary and thyroid as well as in lung and kidney remain unclear, and further series of experiments will be needed to address the local effects of orexins in these tissues.
Interestingly, we found much higher levels of OX1 receptor mRNA in the pituitary and of OX2 receptor mRNA in the adrenal of male compared with female rats. Conversely, hypothalamic OX1 receptor mRNA levels were higher in female rats. Hypothalamic orexin A content is higher in female rats (35), and the effect of orexins on LH release depends on estrogens (12, 13). Thus, sex steroids not only affect the content and actions of orexin, but also may differentially regulate the expression of orexin receptors.
We also show the expression of prepro-orexin mRNA in rat testis. Sakurai et al. indicated the existence of prepro-orexin in rat testis (1). Beside its occurrence in the rat brain, orexin A was found in tissue extracts of rat testis, but not other peripheral tissues (36). Other groups, however, had been unable to detect prepro-orexin (prepro-hypocretin) or orexin A in peripheral rat tissues, including testis (3, 35). These discrepancies may be explained by the different sensitivities of the assays used. Using highly sensitive RT-PCR we were able to detect prepro-orexin mRNA in rat testis. Interestingly, we did not find prepro-orexin mRNA in rat ovaries. Thus, in peripheral tissues orexins appear to be synthesized exclusively in testis. On the other hand, compared with those in brain, prepro-orexin levels in testis were low, and in situ hybridization experiments using oligonucleotide antisense probes failed to reveal significant amounts of prepro-orexin in testis (our unpublished observation). Thus, the functional relevance of testicular prepro-orexin synthesis remains to be clarified.
Prepro-orexin mRNA was not found in other peripheral tissues expressing orexin receptors. Therefore, it is unlikely that in these organs orexins act in an autocrine/paracrine manner. The presence of orexin A in human plasma (37) has been recently described, and we also detected comparable concentrations of orexin A in rat plasma. Thus, circulating orexin may activate orexin receptors in peripheral tissues. The source of plasma orexin remains unclear. As we did not find differences between male and female rats in plasma orexin A levels, it is uncertain whether plasma orexins originate from testis. On the other hand, orexin-containing neurons project to the median eminence and pituitary (32), and orexin may be released into the circulation from the pituitary. As orexin A is relatively slowly metabolized after iv injection (24), a low rate of secretion should be sufficient to contribute to substantial plasma concentrations of orexin. The lack of gender differences in plasma orexin levels and the comparatively low prepro-orexin mRNA expression in testis indicate a predominant role of hypothalamic orexin production. Another possibility to stimulate peripheral orexin receptors could be the action of peptides other than orexin. NPY, for example, has been shown to act at the cloned orexin receptor (38), and NPY is locally produced in the adrenal gland (39).
In summary, we have shown a sexually dimorphic expression of prepro-orexin and orexin receptor subtypes in various peripheral tissues and propose the existence of a peripheral orexin system. Such a system may be involved in the regulation of endocrine and physiological functions that remain to be determined. The localization of OX2 receptor mRNA in the rat adrenal cortex indicates a role in adrenal steroid synthesis and/or release. Our results represent the basis for further physiological studies addressing possible peripheral orexin effects.
|
Acknowledgments |
|---|
|
Footnotes |
|---|
Received December 20, 2000.
Accepted for publication April 3, 2001.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
This article has been cited by other articles:
 |
|
 |
|
  P. Silveyra, V. Lux-Lantos, and C. Libertun Both orexin receptors are expressed in rat ovaries and fluctuate with the estrous cycle: effects of orexin receptor antagonists on gonadotropins and ovulation Am J Physiol Endocrinol Metab, October 1, 2007; 293(4): E977 - E985. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Muller, N. Schweitzer, O. Johren, P. Dominiak, and W. Raasch Angiotensin II stimulates the reactivity of the pituitary-adrenal axis in leptin-resistant Zucker rats, thereby influencing the glucose utilization Am J Physiol Endocrinol Metab, September 1, 2007; 293(3): E802 - E810. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. Silveyra, P. N. Catalano, V. Lux-Lantos, and C. Libertun Impact of proestrous milieu on expression of orexin receptors and prepro-orexin in rat hypothalamus and hypophysis: actions of Cetrorelix and Nembutal Am J Physiol Endocrinol Metab, March 1, 2007; 292(3): E820 - E828. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J E Digby, J Chen, J Y Tang, H Lehnert, R N Matthews, and H S Randeva Orexin receptor expression in human adipose tissue: effects of orexin-A and orexin-B. J. Endocrinol., October 1, 2006; 191(1): 129 - 136. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. Voisin, A. E. Firar, V. Avondo, and M. Laburthe Orexin-Induced Apoptosis: The Key Role of the Seven-Transmembrane Domain Orexin Type 2 Receptor Endocrinology, October 1, 2006; 147(10): 4977 - 4984. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
W. Raasch, C. Wittmershaus, A. Dendorfer, I. Voges, F. Pahlke, C. Dodt, P. Dominiak, and O. Johren Angiotensin II Inhibition Reduces Stress Sensitivity of Hypothalamo-Pituitary-Adrenal Axis in Spontaneously Hypertensive Rats Endocrinology, July 1, 2006; 147(7): 3539 - 3546. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Spinazzi, P. G. Andreis, G. P. Rossi, and G. G. Nussdorfer Orexins in the regulation of the hypothalamic-pituitary-adrenal axis. Pharmacol. Rev., March 1, 2006; 58(1): 46 - 57. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G Sun, Z Tian, Y Yao, H Li, and T Higuchi Central and/or peripheral immunoreactivity of orexin-A in pregnant rats and women J. Mol. Endocrinol., February 1, 2006; 36(1): 131 - 138. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. R. Holland, S. Akerman, and P. J. Goadsby Orexin 1 Receptor Activation Attenuates Neurogenic Dural Vasodilation in an Animal Model of Trigeminovascular Nociception J. Pharmacol. Exp. Ther., December 1, 2005; 315(3): 1380 - 1385. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. L. Barreiro, R. Pineda, F. Gaytan, M. Archanco, M. A. Burrell, J. M. Castellano, H. Hakovirta, M. Nurmio, L. Pinilla, E. Aguilar, et al. Pattern of Orexin Expression and Direct Biological Actions of Orexin-A in Rat Testis Endocrinology, December 1, 2005; 146(12): 5164 - 5175. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Spinazzi, M. Rucinski, G. Neri, L. K. Malendowicz, and G. G. Nussdorfer Preproorexin and Orexin Receptors Are Expressed in Cortisol-Secreting Adrenocortical Adenomas, and Orexins Stimulate in Vitro Cortisol Secretion and Growth of Tumor Cells J. Clin. Endocrinol. Metab., June 1, 2005; 90(6): 3544 - 3549. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Karteris, R. J. Machado, J. Chen, S. Zervou, E. W. Hillhouse, and H. S. Randeva Food deprivation differentially modulates orexin receptor expression and signaling in rat hypothalamus and adrenal cortex Am J Physiol Endocrinol Metab, June 1, 2005; 288(6): E1089 - E1100. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. M. Taylor, J. R. Baker, and W. K. Samson Brain-derived adrenomedullin controls blood volume through the regulation of arginine vasopressin production and release Am J Physiol Regulatory Integrative Comp Physiol, May 1, 2005; 288(5): R1203 - R1210. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Chen and H. S. Randeva Genomic Organization of Mouse Orexin Receptors: Characterization of Two Novel Tissue-Specific Splice Variants Mol. Endocrinol., November 1, 2004; 18(11): 2790 - 2804. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. Rouet-Benzineb, C. Rouyer-Fessard, A. Jarry, V. Avondo, C. Pouzet, M. Yanagisawa, C. Laboisse, M. Laburthe, and T. Voisin Orexins Acting at Native OX1 Receptor in Colon Cancer and Neuroblastoma Cells or at Recombinant OX1 Receptor Suppress Cell Growth by Inducing Apoptosis J. Biol. Chem., October 29, 2004; 279(44): 45875 - 45886. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. W. Kok, F. Roelfsema, S. Overeem, G. J. Lammers, M. Frolich, A. E. Meinders, and H. Pijl Pulsatile LH release is diminished, whereas FSH secretion is normal, in hypocretin-deficient narcoleptic men Am J Physiol Endocrinol Metab, October 1, 2004; 287(4): E630 - E636. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. L. Barreiro, R. Pineda, V. M. Navarro, M. Lopez, J. S. Suominen, L. Pinilla, R. Senaris, J. Toppari, E. Aguilar, C. Dieguez, et al. Orexin 1 Receptor Messenger Ribonucleic Acid Expression and Stimulation of Testosterone Secretion by Orexin-A in Rat Testis Endocrinology, May 1, 2004; 145(5): 2297 - 2306. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Karteris, J. Chen, and H. S. Randeva Expression of Human Prepro-Orexin and Signaling Characteristics of Orexin Receptors in the Male Reproductive System J. Clin. Endocrinol. Metab., April 1, 2004; 89(4): 1957 - 1962. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Ehmke and A. Just The orexins: linking circulatory control with behavior Am J Physiol Regulatory Integrative Comp Physiol, September 1, 2003; 285(3): R519 - R521. [Full Text] [PDF]
|
|
|
|
|
|
O. Johren, N. Bruggemann, A. Dendorfer, and P. Dominiak Gonadal Steroids Differentially Regulate the Messenger Ribonucleic Acid Expression of Pituitary Orexin Type 1 Receptors and Adrenal Orexin Type 2 Receptors Endocrinology, April 1, 2003; 144(4): 1219 - 1225. [Abstract] [Full Text] [PDF]
|
|
|
