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Veterans Affairs Medical Center, Geriatric, Research, Education and Clinical Center and Research Service, Minneapolis 55417; and Department of Food Science and Nutrition, University of Minnesota, St. Paul, Minnesota 55108
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ABSTRACT |
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 TOP
 ABSTRACT
 INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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The intermediate portion of the lateral
septum (LSi) contains high levels of urocortin (UCN) peptide and type 2 corticotropin-releasing hormone (CRH) receptor (CRHR2) and has anatomic
and functional connections with the lateral hypothalamus (LH). We
tested the effect of UCN in the LSi on feeding. Injection of 10 or 30 pmol UCN into LSi significantly decreased feeding in food-deprived rats
for 24 h without producing conditioned taste aversion (CTA). Pretreatment with a CRH receptor antagonist, 
-helical CRH
(-hCRH), blocked the inhibitory effect of UCN on deprivation-induced
feeding at 1 and 2 h postinjection. Furthermore, UCN in the LSi
significantly decreased feeding induced by LH-injected orexin A
at 2 and 4 h postinjection, and addition of -hCRH blocked the
inhibitory effect of UCN on orexin A-induced feeding. In
conclusion, UCN significantly inhibits feeding induced by
deprivation and LH-injected orexin A without producing a CTA, an effect
that is mediated by CRHR2. These data define the LSi as an important
site for UCN-induced anorexia and indicate that LSi UCN may influence
orexin A feeding signals in the LH.
intermediate portion of lateral septum; corticotropin releasing hormone receptor; satiety
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INTRODUCTION |
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TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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UROCORTIN (UCN) is a recently identified member of the corticotropin-releasing hormone (CRH) family (48) that decreases normal and deprivation-induced feeding after intracerebroventricular injection (42). UCN also decreases normal and/or deprivation-induced feeding after injection into the 4th ventricle (19), the hypothalamic paraventricular nucleus (PVN) (49), and the hypothalamic ventromedial nucleus (30, 31) and decreases neuropeptide Y-induced feeding after injection into the PVN (49). UCN binds to all three CRH receptors with higher affinity than CRH but binds with particular avidity to the type 2 CRH receptor (CRHR2) (48). CRHR2 knockout mice and rats treated with antisense to CRHR2 display alterations in feeding behavior (3, 11, 41). Together these studies imply that UCN may be the endogenous ligand for CRHR2 (25, 48). Ventricular injection of UCN decreases feeding with an ED50 25 times less than that for CRH (42) and with attenuated anxiogenic responses compared with that observed after CRH administration. However, the difference in magnitude of feeding inhibition by UCN and CRH in the PVN was not as profound as that observed after intracerebroventricular injection, which suggests the presence of other, potentially more sensitive sites for UCN action.
The lateral septum (LS) is part of the limbic system and is important in memory, learning, reward, stress, and defense mechanisms. The LS may also be important to the regulation of feeding. Lesioning the septum results in increased feeding behavior (24), greater licking responses to sucrose solutions, and exaggerated facilitatory and inhibitory behavior when various solutions (palatable or aversive) or tastants are offered, suggesting that the LS may be involved in the rewarding aspects of feeding (18, 47). Several pieces of evidence suggest that the intermediate portion of the LS (LSi) is an important site of UCN action: UCN has a dense fiber network in the LSi (6, 25); UCN peptide and CRHR2 coexist within the LSi (16, 25, 35, 48); and intracerebroventricular administration of UCN elevates c-Fos expression (an indicator of cellular activation) in CRHR2-containing cells within the LSi (48). Recent data indicate that the LSi is innervated by fibers containing the recently identified UCNIII (also known as stresscopin), suggesting that UCNIII may have important actions in this area (26).
The lateral hypothalamus (LH) is a central feeding center. Electrical stimulation of the LH increases feeding behavior whereas chemical lesions in this region result in anorexia and death (5). There is a predominance of orexin A (also referred to as hypocretin 1) and orexin receptors within the LH (7, 14, 40), and injection of orexin A into the LH induces feeding (15, 43). Neuroanatomic tracing studies indicate that the LS receives neural inputs from the LH and sends monosynaptic projections to the LH (37), which provides an anatomic basis for LS-LH signaling. Several studies also demonstrate a functional connection between the LS and the LH in the regulation of feeding. Electrical stimulation of the LS significantly inhibits feeding induced by stimulation of the LH, whereas electrolytic damage of the LS enhances feeding induced by stimulation of the LH, implicating inhibitory control of the LH by the LS (32). The recent demonstration that electrical stimulation of the mediolateral portion of the LS (LSml, adjacent to the LSi) results in elevated c-Fos immunoreactivity in the LH further suggests that these two regions are functionally connected (46). Another recent study indicates that ventricular administration of UCN and CRH results in elevated thresholds for LH self-stimulation, suggesting that UCN and/or CRH may modulate LH reward processes (27).
On the basis of the above findings, we hypothesized that UCN within the LSi inhibits feeding induced by deprivation and LH orexin A stimulation and that UCN effects within the LSi are mediated through CRHR2. We performed a series of experiments to test these hypotheses, and our studies show that 1) UCN in the LSi inhibits deprivation-induced feeding without producing a conditioned taste aversion (CTA); 2) UCN in the LSi inhibits feeding induced by orexin A in the LH; and 3) antagonism of CRH receptor blocks the inhibitory effect of LSi-injected UCN on feeding induced by food deprivation and orexin A in the LH.
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METHODS |
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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The treatment of animals in these studies fully conforms with the Guiding Principles for Research Involving Animals and Human Beings of the American Physiological Society (1), and these studies received local institutional animal care and use committee approval.
Animals
Male Sprague-Dawley rats (Harlan, Madison, WI) weighing 280-325 g were housed individually in cages with a 12:12-h light-dark photocycle (lights on at 0700) in a room at 21-22°C. Teklad lab chow and water were allowed ad libitum, except where noted.Cannulation and Verification of Placement
Rats were anesthetized with Nembutal (40 mg/kg) and were fitted with a 28-gauge stainless steel guide cannula placed just above the LSi and/or LH. Stereotaxic coordinates for the LSi were determined from the rat brain atlas by Paxinos and Watson (33) and were as follows: 0.4 mm lateral and 0.8 mm anterior to bregma, and 4.8 mm below the skull surface. For experiments 1a and 2-5, injections were unilateral on the right side. For experiment 1b, injections were bilateral into the LSi. Stereotaxic coordinates for the LH were 2.0 mm lateral and 2.1 mm posterior to bregma, and 7.3 mm below the skull surface. These coordinates were based on our previous studies indicating that this region of the LH is sensitive to the feeding effects of orexin A (43). Injectors extended 1 mm beyond the end of the guide cannula. The animals were given at least 1 wk to recover after surgery before experimental trials. After the experiments brains were dissected out and stored in a 10% formaldehyde solution for histological placement verification. A cannula was deemed incorrect if the injection site was outside of the region targeted and further than 0.75 mm away from the targeted site. Most injections (~85%) occurred within a 0.5-mm radius from the targeted site. Data from animals with incorrectly placed cannulas were excluded from the final analysis. The number of rats listed in the specific experimental protocols represents the number of rats in the final analysis (all cannulas correctly placed). Figure 1 illustrates examples of histological verification of correct location of injections into the LSi and into the LH. Figure 2 illustrates a map of actual injection sites and demonstrates those injections deemed correctly and incorrectly located for one study (experiment 5). This map is representative of placement verification for all studies and represents the widest possible variation in injection sites for all studies.
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Drugs
UCN and orexin A were purchased from Phoenix Pharmaceuticals (Mountain View, CA), and-helical CRH (-hCRH) was purchased from
Sigma (St. Louis, MO). These compounds were dissolved in artificial
cerebrospinal fluid (aCSF) just before use.
Injections
A volume of 0.5 µl was injected slowly over 30 s, with injector left in place an additional 10 s to ensure extrusion from the tip and to minimize distribution of drug upward on the cannula tract. The total number of injections for each animal was <12. In previous studies, we demonstrated a lack of extensive tissue damage after 50 repeated injections as measured by gliosis around the injection site (10) and light microscopy at ×100. Injection sites were examined by light microscopy for extensive tissue damage in the present studies and none was found.Specific Experimental Protocols
Experiment 1a: effect of unilateral injections of UCN and CRH in the LSi on deprivation-induced feeding. Seven LSi-cannulated rats were food deprived for 18 h and injected with 0 (vehicle = aCSF), 3, 10, or 30 pmol UCN. Food was given immediately after injection, and food intake was measured at 1, 2, 4, and 24 h after injection. Body weight was measured at 24 h after injection. Each animal received each treatment once with at least 72 h between treatments to allow for clearance of drug from the central nervous system (CNS) and for normal feeding patterns to be reestablished. Treatments were given in a randomly selected counterbalanced design. After a 1-wk washout period after the first feeding study, these animals were injected with the same doses of CRH under the same protocol.
Experiment 1b: comparison between unilateral vs. bilateral injections of UCN in the LSi on deprivation-induced feeding. Nine bilaterally LSi-cannulated rats were food deprived for 18 h and injected with 0 (vehicle = aCSF), 10 or 30 pmol UCN unilaerally or bilaterally. For the bilateral injection, 10 or 30 pmol UCN were injected on each side and thus the total dose was 20 or 60 pmol. Food was given immediately after injection, and food intake was measured at 1, 2, 4, and 24 h after injection. Body weight was measured at 24 h after injection. Each animal received each treatment once with at least 72 h between treatments to allow for clearance of drug from the CNS and for normal feeding patterns to be reestablished. Treatments were given in a randomly selected counterbalanced design.
Experiment 2: CTA. The two-bottle preference test was used to determine whether UCN is aversive when administered into the LSi. The basis for this is as follows. When rats are exposed to water and saccharin solutions they show a preference for saccharin. When animals are exposed to saccharin for the first time and concurrently injected with a drug that has aversive properties, saccharin is associated with the aversive stimuli. Subsequently, when given water and saccharin at the same time, drinking of saccharin is reduced. Reduced consumption of the drug-paired flavor indicates that the drug administered has aversive properties. Twenty rats were water deprived for 23.5 h and had water access for 0.5-h each day for 7 days. They were then randomly divided into four groups (with an even distribution of body weight) and given 15 ml of saccharin immediately followed by injection with 0 (vehicle = aCSF), or 10, 30, or 100 pmol UCN. This conditioned stimulation was repeated once after 2 days, and the animals were then given the choice of water and saccharin in the absence of drug administration 2 days later. Twentyfour-hour intake of water and saccharin was measured.
Experiment 3: pretreatment of the LSi with CRH receptor
antagonist: effect on LSi UCN-induced feeding inhibition.
Nine LSi-cannulated rats were food-deprived for 18 h and randomly
assigned to four treatments: 1) aCSF + aCSF,
2) aCSF + UCN (30 pmol), 3) -hCRH (1 µg) + UCN (30 pmol), or 4) -hCRH (1 µg) + aCSF. The first injection was given 15 min before the second injection.
Food was given immediately after the second injection. The dose of 30 pmol of UCN was chosen based on its inhibitory effect on feeding in the
absence of a CTA in experiment 2. The dose of -hCRH was
based on published doses used for intracerebroventricular (28) and PVN (20) administration. Each animal
received each treatment once with at least 72 h between treatments.
Experiment 4: effect of UCN in the LSi on feeding induced by orexin A in the LH. Eighteen nondeprived LSi and LH double-cannulated rats were randomly assigned to one of four treatments: 1) aCSF + aCSF, 2) aCSF + orexin A (1,000 pmol), 3) UCN (30 pmol) + orexin A (1,000 pmol), or 4) UCN (30 pmol) + aCSF. Animals were given the first injection 15 min before the second injection. Injections were carried out during the light cycle between 1230 and 1400. Each animal received each treatment once with at least 72 h between treatments.
Experiment 5: pretreatment of the LSi with a CRH receptor
antagonist: effect on LSi-injected UCN inhibition of feeding produced
by LH-injected orexin A.
Eight nondeprived LSi and LH double-cannulated rats were randomly
assigned to one of four treatments: 1) LSi (aCSF + aCSF) + LH (aCSF), 2) LSi (aCSF + aCSF) + LH
(orexin A, 1,000 pmol), 3) LSi (aCSF + UCN, 30 pmol) + LH (orexin A, 1,000 pmol), or 4) LSi [-hCRH
(1 µg) + UCN (30 pmol)] + LH (aCSF). The first LSi injection
was given 15 s before the second LSi injection. The LH injection
was given 15 min after the second LSi injection. Food intake was
measured at 1, 2, and 4 h after the LH injection. Injections were
carried out during the light cycle between 1230 and 1400. Each animal
received each treatment once with at least 72 h between treatments.
Statistical Analyses
For experiments 1a and 3-5, the data were analyzed by repeated-measures ANOVA, and thus each rat served as its own control. When main effects were observed, post hoc analysis was performed using multiple-comparison contrasts. For experiment 1b, data were analyzed by a two-factor ANOVA followed by Fisher's least-significant difference t-test to compare means. For experiment 2, data were analyzed by a one-factor ANOVA followed by Fisher's least-significant difference t-test to compare means.| |
RESULTS |
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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Experiment 1a: Effect of LSi-injected UCN and CRH on Deprivation-Induced Feeding
In the first hour, LSi-injected UCN at 3, 10, and 30 pmol significantly inhibited deprivation-induced feeding by 20.6% (P = 0.015), 42.8% (P < 0.0001), and 49.4% (P < 0.0001), respectively (Fig. 3). Two hours after injection, these same doses of UCN inhibited feeding by 22.8% (P = 0.0203), 24.4% (P = 0.014), and 44.3% (P < 0.0001), respectively (Fig. 3). At 24 h postinjection, UCN at 10 and 30 pmol significantly inhibited feeding by 9.1% (P = 0.0358) and 10.6% (P = 0.0163), respectively (Fig. 3). UCN at 10 and 30 pmol significantly reduced body weight gain by 24.4% (P = 0.0037) and 17.3% (P = 0.0307), respectively (Fig. 5).
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LSi-injected CRH at 3, 10, and 30 pmol significantly inhibited feeding
by 29.9% (P = 0.0053), 48.7% (P < 0.0001), and 48.9% (P < 0.0001), respectively, at
1 h after injection (Fig. 4).
However, there was no significant feeding inhibition after the first
hour, and CRH did not significantly influence body weight gain measured 24 h after injection (Fig. 5).
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Experiment 1b: Effect of Unilateral vs. Bilateral Injection of UCN in LSi on Deprivation-Induced Feeding
Two-factor ANOVA, with injection type (bilateral or unilateral) and UCN doses representing factors, indicates no significant difference in feeding response to unilateral and bilateral injection of UCN at 1, 4, and 24 h after injection (P = 0.503, P = 0.8593, and P = 0.5899, respectively; Fig. 6). However, there was a main effect of UCN treatment at 1, 4, and 24 h after injection (P = 0.0078, P = 0.0126, and P = 0.0078, respectively).
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Experiment 2: Effect of LSi-Injected UCN on Preference for Saccharin Solution
There were main effects of UCN on the percentage of fluid intake attributed to saccharin solution. Intake of saccharin solution in the group treated with 100 pmol UCN was significantly lower than that in the animals treated with aCSF (P = 0.0092, Fig. 7). There was no significant difference in saccharin solution intake among the groups receiving 0, 10, and 30 pmol of UCN (Fig. 7).
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Experiment 3: Pretreatment of the LSi with CRH Receptor Antagonist and Effect on LSi UCN-Induced Feeding Inhibition
In this experiment,-hCRH was injected into the LSi 15 min
before LSi injection of UCN in food-deprived animals. In the first hour
after injection, UCN at 30 pmol significantly inhibited
deprivation-induced feeding by 29.3% (P = 0.0007, Fig.
8) compared with control levels, and
addition of 1 µg -hCRH significantly blocked the inhibitory effect
of UCN (P = 0.0006, Fig. 8), resulting in a food intake value that was 42.4% greater than the level observed after UCN treatment (aCSF + UCN). At 2 h after injection, UCN decreased feeding by 26.3% (P = 0.0034, Fig. 8), and addition of
-hCRH blocked the anorectic effect of UCN, increasing feeding by
31.3% (P = 0.0088, Fig. 8) compared with UCN treatment
alone (aCSF + UCN). -hCRH treatment alone (-hCRH + aCSF) did not increase feeding above control levels at any time
point.
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Experiment 4: Effect of LSi-Injected UCN on Feeding Induced by LH-Injected Orexin A
In the first hour postinjection, orexin A administered into the LH significantly increased feeding by 3.2-fold above control levels (P = 0.0001, Fig. 9), and UCN at 30 pmol did not significantly block orexin A-induced feeding (Fig. 9). In the second hour after injection, UCN significantly inhibited orexin A-induced feeding by 64.3% (P = 0.0208, Fig. 9). Thus within 2 h postinjection, LSi UCN blocked LH orexin A-induced feeding by 30.9% (P = 0.057, Fig. 9). At 4 h postinjection, orexin A insignificantly increased feeding, and UCN blocked LH orexin A-induced feeding by 34.7% (P = 0.028), resulting in a level of food intake that was not significantly different from controls (Fig. 9).
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Experiment 5: Pretreatment of the LSi with a CRH Receptor Antagonist and Effect on LSi-Injected UCN Inhibition of Feeding Produced by LH-Injected Orexin A
In the first hour, orexin A in the LH significantly increased feeding by 2.1-fold above control levels (P = 0.005), and LSi-injected UCN blocked LH orexin A-induced feeding by 56.2% (P = 0.0156, Fig. 10). However, addition of-hCRH into the LSi did not block the anorectic
effect of UCN. By 2 and 4 h postinjection, -hCRH in the LSi
significantly blocked the inhibitory effect of LSi UCN on LH-orexin
A-induced feeding (P = 0.0426 and P = 0.0047, respectively, Fig. 10).
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DISCUSSION |
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TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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The current data demonstrate for the first time that UCN acts within the LSi to inhibit feeding behavior. Previously there had been no identified neurochemical basis for the anorectic response to stimulation of the LS (32). Our studies show that UCN in the LSi inhibits deprivation-induced feeding without producing a CTA, inhibits feeding induced by orexin A in the LH, and mediates feeding inhibition by CRH receptor stimulation. These findings are consistent with the strong neurochemical and neuroanatomic basis for UCN action within the LSI: UCN peptide is heavily distributed in the LSi and matches the presence of CRHR2 in the LSi. Moreover, intracerebroventricular administration of UCN activates c-Fos expression in CRHR2-containing cells within the LSi (48). The LS area lines the lateral cerebroventricles and may be relatively accessible to cerebrospinal fluid components and ventricular injectates. It is possible that the profound feeding-inhibitory effects observed with lateral ventricular injection of UCN and/or CRH represent actions within the LSi. Furthermore, the LS had been previously implicated in feeding behavior by lesioning and electrical stimulation studies: lesions of the LS resulted in stimulation of feeding (24) and increased sensitivity to specific tastants (18, 47), whereas electrical stimulation of the LS decreased feeding (24). As part of the limbic system the LS may relay signals about sensory and reward properties of feeding, and due to the large hippocampal input to the LS there may also be an association with the learning and memory aspects of feeding (44).
Unilateral administration of UCN in the LSi decreased feeding induced by deprivation (Fig. 3), which was associated with changes in body weight (Fig. 5) and is consistent with studies demonstrating that electrical stimulation of LSi inhibits feeding (24). Bilateral injection of UCN did not result in further reduction of deprivation-induced feeding (Fig. 6), indicating that the level of reduction observed with unilateral injection is the maximal response. This lack of complete abolition of the feeding response to food deprivation is not surprising given the multitude of feeding signals generated in response to food deprivation. LSi-injected CRH also inhibited deprivation-induced feeding (Fig. 4). However, the feeding inhibition by CRH lasted only 1 h, much shorter than the 24-h feeding inhibition observed after UCN injection, which is consistent with the higher affinity of UCN for CRHR2 than that of CRH for CRHR2 (48). The enhanced efficacy of LSi-injected UCN vs. CRH in feeding inhibition (Figs. 3-5) and the heavy distribution of UCN peptide in the LSi suggests that UCN, rather than CRH, is the endogenous ligand for CRHR2 in the LSi. Involvement of CRHR2 in feeding is in agreement with studies in which both CRHR2 knockout mice and rats treated with CRHR2 antisense or other CRH-selective antagonists (antisauvagine-30) display alterations in feeding behavior (3, 11, 12, 22, 41).
To verify that the feeding inhibition observed after LSi-injected UCN is not due to potential aversive effects of UCN, a CTA study was performed. UCN at doses equal and less than 30 pmol, doses for which significant feeding inhibition is observed, did not induce taste aversion, and this indicates that feeding inhibition by these doses of UCN is not due to malaise or other aversive sequelae (Fig. 7). Animals conditioned to associate saccharin consumption with a high dose of UCN (100 pmol) significantly reduced consumption of saccharin in subsequent exposures, indicating that saccharin had become aversive as a result of high-dose UCN administration (Fig. 7).
To establish that feeding inhibition induced by LSi-injected UCN is
mediated via CRHR2, we tested the effect of -hCRH on feeding
inhibition produced by UCN administration in the LSi. -hCRH is a
synthetic antagonist of CRH receptor and binds to both the type 1 CRH
receptor (CRHR1) and to CRHR2. Because CRHR2 is the only CRH receptor
subtype within the LSi (6, 8, 45), -hCRH
injected in the LSi would bind to CRHR2 exclusively. As shown in Fig.
8, -hCRH blocks UCN inhibition of deprivation-induced feeding,
suggesting that UCN action in the LSi is mediated by CRHR2. However, as
demonstrated in Fig. 8, blockade of CRHR2 in the LSi without concurrent
UCN treatment did not enhance deprivation-induced feeding. It is
possible that blockade of CRHR2 in the LSi in nondeprived rats may
enhance normal feeding, but this was not tested in the current study.
We also sought to determine whether UCN in the LSi influences feeding signals generated by stimulation of the LH with orexin A. Orexin-containing neurons in the LH project throughout the neuroaxis (34), and the two receptors for the orexins, type 1 (OX1R) and type 2 (OX2R), are also widely distributed. Orexin A has a high affinity for OX1R (39, 40), and recent evidence indicates that OX1R is present in the LH (2). The neural mechanism by which LH-injected orexin A stimulates feeding is unknown. In the LH, OX1R has been demonstrated in cells containing melanin-concentrating hormone (MCH), which coexpress cocaine-amphetamine related transcript (17), and in cells expressing orexin (2), which coexpress prodynorphin (9). A recent study indicates that orexin may interact with MCH-containing neurons (4), and it is possible that orexin stimulation of the LH enhances feeding via activation of MCH-containing neurons, as MCH elicits feeding on central administration (36). However, the presence of several feeding-related neuropeptides in the LH and the wide distribution of orexin-containing neurons throughout the brain in areas implicated in a variety of functions suggest that the stimulation of feeding by orexin A in the LH likely involves multiple mechanisms.
Several studies suggest communication between the LS area and the LH in
the regulation of feeding behavior. The LSi sends monosynaptic
projections to the LH (37), and the LH has a
well-established role in feeding behavior (38). Lesions of
the LS enhance LH-stimulated feeding behavior, whereas stimulation of
the LS inhibits LH-stimulated feeding (32). Furthermore, a
recent demonstration that electrical stimulation of the LSml (adjacent
to the LSi) results in elevated c-Fos immunoreactivity in the LH
suggests that these two regions are functionally connected
(46). A recent study showing elevated thresholds for LH
self-stimulation after ventricular administration of UCN and CRH also
suggests that UCN and/or CRH may modulate LH reward processes
(27). On the basis of these anatomic and functional
connections between the LSi and LH, we hypothesized that UCN may
decrease feeding by influencing LH-induced feeding signals. As
demonstrated in Fig. 9, orexin A in the LH significantly increased
feeding, and this increase was blocked by injection of UCN in the LSi.
Furthermore, we found that addition of -CRH in the LSi before UCN
injection blocked the inhibitory effect of UCN, and food intake in the
animals treated with -hCRH, UCN, and orexin A was similar for
animals treated with LH-orexin A alone (Fig. 10). Although the specific
neurons activated by UCN in the LS are unknown, there are several cell
types present in the LS region, including gonadotropin-releasing
hormone (13), GABAergic calbindin cells, orphanin FQ
(21, 29), somatostatin, and neurotensin (23),
and it is possible that stimulation of CRHR2 influences some of these
neurons. However, whether these specific neurons project to the LH
region we targeted is unknown.
These data suggest that when signaling mechanisms in the LSi are
increased (via UCN), feeding response to LH stimulation (by orexin A)
is decreased, suggesting that UCN stimulation of the LSi transmits
feeding inhibitory signals to the LH. Conversely, when signaling
mechanisms in the LSi are decreased (via -hCRH), feeding
response to LH stimulation (by orexin A) is increased, suggesting that
communication of inhibitory messages received by the LH are prevented.
These data agree with reports by Oliveira et al. (32), who
demonstrated that when signaling in the LSi is increased by electrical
stimulation, feeding response to LH stimulation is decreased.
Conversely, when signaling in the LSi was decreased by permanent
lesions, the feeding response to LH stimulation was increased
(32). Together, these data suggest LSi inhibition of LH
feeding signals and also implicate UCN and orexin A as neuropeptides
that permit communication between the LSi and the LH in the control of feeding.
There are some inconsistencies related to the timing of feeding
inhibition in these studies. In experiment 3, reversal of UCN-induced inhibition of deprivation-induced feeding by -hCRH occurred in the first hour (Fig. 8), whereas in experiment
5, the effect of -hCRH on UCN inhibition of LH orexin A-induced feeding only occurred after 1 h (Fig. 10). The second discrepancy relates to the feeding-inhibitory effect of LSi-injected UCN. In most
experiments, UCN was effective in decreasing feeding within the first
hour (Figs. 3, 6, and 8). However, in one experiment, the effect was
significant only after hour 1 (Fig. 9). Although there
is no clear explanation for these differences, variations in strength
of feeding stimulus (deprivation vs. orexin A stimulation), random day
effects, and unknown between-lot differences in efficacy of peptides
are potential explanations that may account for some of the variability observed.
In conclusion, UCN in the LSi inhibits deprivation-induced feeding without producing a CTA, and the inhibitory effect lasts up to 24 h. Furthermore, UCN in the LSi inhibits feeding induced by orexin A in the LH, suggesting alteration of signaling mechanisms between the LSi and the LH. Finally, the inhibitory effect of UCN in the LSi on deprivation- and orexin A-induced feeding is blocked by antagonism of CRH receptor. Together, these data define the LSi region as an important site for UCN-induced anorexia mediated by CRHR2 and suggest that LSi UCN may influence orexin A feeding signals in the LH.
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FOOTNOTES |
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Address for reprint requests and other correspondence: C. M. Kotz, Veterans Affairs Medical Center, GRECC (11G), One Veterans Drive, Minneapolis, MN 55417 (E-mail: kotzx004{at}umn.edu).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
10.1152/ajpregu.00558.2001
Received 17 September 2001; accepted in final form 2 May 2002.
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REFERENCES |
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TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES
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1.
American Physiological Society.
Guiding principles for research involving animals and human beings.
Am J Physiol Regul Integr Comp Physiol
283:
R281-R283,
2002
2.
Backberg, M,
Hervieu G,
Wilson S,
and
Meister B.
Orexin receptor-1 (OX-R1) immunoreactivity in chemically identified neurons of the hypothalamus: focus on orexin targets involved in control of food and water intake.
Eur J Neurosci
15:
315-328,
2002[ISI][Medline].
3.
Bale, TL,
Contarino A,
Smith GW,
Chan R,
Gold LH,
Sawchenko PE,
Koob GF,
Vale WW,
and
Lee KF.
Mice deficient for corticotropin-releasing hormone receptor-2 display anxiety-like behaviour and are hypersensitive to stress.
Nat Genet
24:
410-414,
2000[ISI][Medline].
4.
Bayer, L,
Mairet-Coello G,
Risold PY,
and
Griffond B.
Orexin/hypocretin neurons: chemical phenotype and possible interactions with melanin-concentrating hormone neurons.
Regul Pept
104:
33-39,
2002[ISI][Medline].
5.
Bernardis, LL,
and
Bellinger LL.
The lateral hypothalamic area revisited: ingestive behavior.
Neurosci Biobehav Rev
20:
189-287,
1996[ISI][Medline].
6.
Bittencourt, JC,
Vaughan J,
Arias C,
Rissman RA,
Vale WW,
and
Sawchenko PE.
Urocortin expression in rat brain: evidence against a pervasive relationship of urocortin-containing projections with targets bearing type 2 CRF receptors.
J Comp Neurol
415:
285-312,
1999[ISI][Medline].
7.
Broberger, C,
De Lecea L,
Sutcliffe JG,
and
Hokfelt T.
Hypocretin/orexin- and melanin-concentrating hormone-expressing cells form distinct populations in the rodent lateral hypothalamus: relationship to the neuropeptide Y and agouti gene-related protein systems.
J Comp Neurol
402:
460-474,
1998[ISI][Medline].
8.
Chalmers, DT,
Lovenberg TW,
and
De Souza EB.
Localization of novel corticotropin-releasing factor receptor (CRF2) mRNA expression to specific subcortical nuclei in rat brain: comparison with CRF1 receptor mRNA expression.
J Neurosci
15:
6340-6350,
1995
9.
Chou, TC,
Lee CE,
Lu J,
Elmquist JK,
Hara J,
Willie JT,
Beuckmann CT,
Chemelli RM,
Sakurai T,
Yanagisawa M,
Saper CB,
and
Scammell TE.
Orexin (hypocretin) neurons contain dynorphin.
J Neurosci
21:
RC168,
2001
10.
Cleary, JP,
O'Hare E,
Pomonis JD,
Dittel PL,
Hofmeister JJ,
Fritz MM,
Billington CJ,
and
Levine AS.
Discriminative stimulus effects of morphine: central versus peripheral training.
Brain Res
847:
26-31,
1999[ISI][Medline].
11.
Coste, SC,
Kesterson RA,
Heldwein KA,
Stevens SL,
Heard AD,
Hollis JH,
Murray SE,
Hill JK,
Pantely GA,
Hohimer AR,
Hatton DC,
Phillips TJ,
Finn DA,
Low MJ,
Rittenberg MB,
Stenzel P,
and
Stenzel-Poore M. P.
Abnormal adaptations to stress and impaired cardiovascular function in mice lacking corticotropin-releasing hormone receptor-2.
Nat Genet
24:
403-409,
2000[ISI][Medline].
12.
Cullen, MJ,
Ling N,
Foster AC,
and
Pelleymounter MA.
Urocortin, corticotropin releasing factor-2 receptors and energy balance.
Endocrinology
142:
992-999,
2001
13.
Daikoku, S,
Hisano S,
and
Maki Y.
Immunohistochemical demonstration of LHRH-neurons in young rat hypothalamus: light and electron microscopy.
Arch Histol Jpn
45:
69-82,
1982[Medline].
14.
De Lecea, L,
Kilduff TS,
Peyron C,
Gao X,
Foye PE,
Danielson PE,
Fukuhara C,
Battenberg EL,
Gautvik VT,
Bartlett FS,
Frankel WN,
van den Pol AN,
Bloom FE,
Gautvik KM,
and
Sutcliffe JG.
The hypocretins: hypothalamus-specific peptides with neuroexcitatory activity.
Proc Natl Acad Sci USA
95:
322-327,
1998
15.
Dube, MG,
Kalra SP,
and
Kalra PS.
Food intake elicited by central administration of orexins/hypocretins: identification of hypothalamic sites of action.
Brain Res
842:
473-477,
1999[ISI][Medline].
16.
Eckart, K,
Radulovic J,
Radulovic M,
Jahn O,
Blank T,
Stiedl O,
and
Spiess J.
Actions of CRF and its analogs.
Curr Med Chem
6:
1035-1053,
1999[ISI][Medline].
17.
Elias, CF,
Lee CE,
Kelly JF,
Ahima RS,
Kuhar M,
Saper CB,
and
Elmquist JK.
Characterization of CART neurons in the rat and human hypothalamus.
J Comp Neurol
432:
1-19,
2001[ISI][Medline].
18.
Fried, PA.
Septum and behavior: a review.
Psychol Bull
78:
292-310,
1972[ISI][Medline].
19.
Grill, HJ,
Markison S,
Ginsberg A,
and
Kaplan JM.
Long-term effects on feeding and body weight after stimulation of forebrain or hindbrain CRH receptors with urocortin.
Brain Res
867:
19-28,
2000[ISI][Medline].
20.
Heinrichs, SC,
Menazghi G,
Merlo Pich E,
Hauger RL,
and
Koob GF.
Corticotropin-releasing factor in the paraventricular nucleus modulates feeding induced by neuropeptide Y.
Brain Res
611:
18-24,
1993[ISI][Medline].
21.
Ikeda, K,
Watanabe M,
Ichikawa T,
Kobayashi T,
Yano R,
and
Kumanishi T.
Distribution of prepro-nociceptin/orphanin FQ mRNA and its receptor mRNA in developing and adult mouse central nervous systems.
J Comp Neurol
399:
139-151,
1998[ISI][Medline].
22.
Kishimoto, T,
Radulovic J,
Radulovic M,
Lin CR,
Schrick C,
Hooshmand F,
Hermanson O,
Rosenfeld MG,
and
Spiess J.
Deletion of crhr2 reveals an anxiolytic role for corticotropin-releasing hormone receptor-2.
Nat Genet
24:
415-419,
2000[ISI][Medline].
23.
Kohler, C,
and
Eriksson LG.
An immunohistochemical study of somatostatin and neurotensin positive neurons in the septal nuclei of the rat brain.
Anat Embryol (Berl)
170:
1-10,
1984[Medline].
24.
Koppell, S,
and
Sodetz FJ.
Septal ablation in the rat and bar pressing under appetitive and aversive control.
J Comp Physiol Psychol
81:
274-280,
1972[ISI][Medline].
25.
Kozicz, T,
Yanaihara H,
and
Arimura A.
Distribution of urocortin-like immunoreactivity in the central nervous system of the rat.
J Comp Neurol
391:
1-10,
1998[ISI][Medline].
26.
Li, C,
Vaughan J,
Sawchenko PE,
and
Vale WW.
Urocortin III-immunoreactive projections in rat brain: partial overlap with sites of type 2 corticotrophin-releasing factor receptor expression.
J Neurosci
22:
991-1001,
2002
27.
Macey, DJ,
Koob GF,
and
Markou A.
CRF and urocortin decreased brain stimulation reward in the rat: reversal by a CRF receptor antagonist.
Brain Res
866:
82-91,
2000[ISI][Medline].
28.
Monnikes, H,
Schmidt BG,
Raybould HE,
and
Tache Y.
CRF in the paraventricular nucleus mediates gastric and colonic motor response to restraint stress.
Am J Physiol Gastrointest Liver Physiol
262:
G137-G143,
1992
29.
Neal, CR, Jr,
Mansour A,
Reinscheid R,
Nothacker HP,
Civelli O,
and
Watson SJ, Jr.
Localization of orphanin FQ (nociceptin) peptide and messenger RNA in the central nervous system of the rat.
J Comp Neurol
406:
503-547,
1999[ISI][Medline].
30.
Ohata H, Suzuki K, Oki Y, Arai K, and Sjibasaki T. Urocortin in
the ventromedial nucleus of the hypothalamus (VMH) inhibits feeding
behavior in rats (Abstract). Proc 80th Annual Mtg Endocr Soc New
Orleans, LA OR22-2: 95, 1998.
31.
Ohata, H,
Suzuki K,
Oki Y,
and
Shibasaki T.
Urocortin in the ventromedial hypothalamic nucleus acts as an inhibitor of feeding behavior in rats.
Brain Res
861:
1-7,
2000[ISI][Medline].
32.
Oliveira, LA,
Gentil CG,
and
Covian MR.
Role of the septal area in feeding behavior elicited by electrical stimulation of the lateral hypothalamus of the rat.
Braz J Med Biol Res
23:
49-58,
1990[ISI][Medline].
33.
Paxinos, G,
and
Watson C.
The Rat Brain in Stereotaxic Coordinates. San Diego, CA: Academic, 1997.
34.
Peyron, C,
Tighe DK,
van den Pol AN,
de Lecea L,
Heller HC,
Sutcliffe JG,
and
Kilduff TS.
Neurons containing hypocretin (orexin) project to multiple neuronal systems.
J Neurosci
18:
9996-10015,
1998
35.
Primus, RJ,
Yevich E,
Baltazar C,
and
Gallager DW.
Autoradiographic localization of CRF1 and CRF2 binding sites in adult rat brain.
Neuropsychopharmacology
17:
308-316,
1997[ISI][Medline].
36.
Qu, D,
Ludwig DS,
Gammeltoft S,
Piper M,
Pelleymounter MA,
Cullen MJ,
Mathes WF,
Przypek R,
Kanarek R,
and
Maratos-Flier E.
A role for melanin-concentrating hormone in the central regulation of feeding behaviour.
Nature
380:
243-247,
1996[Medline].
37.
Risold, PY,
and
Swanson LW.
Connections of the rat lateral septal complex.
Brain Res Rev
24:
115-195,
1997[Medline].
38.
Rolls, ET.
The neurophysiology of feeding.
Int J Obes
8:
139-150,
1984.
39.
Sakurai, T.
Orexins and orexin receptors: implication in feeding behavior.
Regul Pept
85:
25-30,
1999[ISI][Medline].
40.
Sakurai, T,
Amemiya A,
Ishii M,
Matsuzaki I,
Chemelli RM,
Tanaka H,
Williams SC,
Richardson JA,
Kozlowski GP,
Wilson S,
Arch JR,
Buckingham RE,
Haynes AC,
Carr SA,
Annan RS,
McNulty DE,
Liu WS,
Terrett JA,
Elshourbagy NA,
Bergsma DJ,
and
Yanagisawa M.
Orexins and orexin receptors: a family of hypothalamic neuropeptides and G protein-coupled receptors that regulate feeding behavior.
Cell
92:
573-585,
1998[ISI][Medline].
41.
Smagin, GN,
Howell LA,
Ryan DH,
De Souza EB,
and
Harris RB.
The role of CRF2 receptors in corticotropin-releasing factor- and urocortin-induced anorexia.
Neuroreport
9:
1601-1606,
1998[ISI][Medline].
42.
Spina, M,
Merlo-Pich E,
Chan RK,
Basso AM,
Rivier J,
Vale W,
and
Koob GF.
Appetite-suppressing effects of urocortin, a CRF-related neuropeptide.
Science
273:
1561-1564,
1996[Abstract].
43.
Sweet, DC,
Levine AS,
Billington CJ,
and
Kotz CM.
Feeding response to central orexins.
Brain Res
821:
535-538,
1999[ISI][Medline].
44.
Urban, IJ.
Effects of vasopressin and related peptides on neurons of the rat lateral septum and ventral hippocampus.
Prog Brain Res
119:
285-310,
1998[ISI][Medline].
45.
Van Pett, K,
Viau V,
Bittencourt JC,
Chan RK,
Li HY,
Arias C,
Prins GS,
Perrin M,
Vale W,
and
Sawchenko PE.
Distribution of mRNAs encoding CRF receptors in brain and pituitary of rat and mouse.
J Comp Neurol
428:
191-212,
2000[ISI][Medline].
46.
Varoqueaux, F,
and
Poulain P.
Projections of the mediolateral part of the lateral septum to the hypothalamus, revealed by Fos expression and axonal tracing in rats.
Anat Embryol (Berl)
199:
249-263,
1998.
47.
Vasudev, R,
Gentil CG,
and
Covian MR.
Taste preference in a free-choice situation following electrical stimulation and lesion of septal area in rats.
Physiol Behav
34:
619-624,
1985[Medline].
48.
Vaughan, J,
Donaldson C,
Bittencourt J,
Perrin MH,
Lewis K,
Sutton S,
Chan R,
Turnbull AV,
Lovejoy D,
and
Rivier C.
Urocortin, a mammalian neuropeptide related to fish urotensin I and to corticotropin-releasing factor.
Nature
378:
287-292,
1995[Medline].
49.
Wang, C,
Mullet MA,
Glass MJ,
Billington CJ,
Levine AS,
and
Kotz CM.
Feeding inhibition by urocortin in the rat hypothalamic paraventricular nucleus.
Am J Physiol Regul Integr Comp Physiol
280:
R473-R480,
2001
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