AJP - Regu Watch the video to learn how APS reaches out to developing nations.
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Am J Physiol Regul Integr Comp Physiol 286: R303-R310, 2004; doi:10.1152/ajpregu.00504.2003
0363-6119/04 $5.00
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Web of Science (11)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Zhang, Z.-H.
Right arrow Articles by Felder, R. B.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Zhang, Z.-H.
Right arrow Articles by Felder, R. B.

APPETITE, OBESITY AND METABOLISM

Melanocortin receptors mediate the excitatory effects of blood-borne murine leptin on hypothalamic paraventricular neurons in rat

Zhi-Hua Zhang1 and Robert B. Felder1,2

1Department of Internal Medicine, University of Iowa; and 2Veterans Affairs Medical Center, Iowa City, Iowa 52242

Submitted 2 September 2003 ; accepted in final form 25 October 2003


   ABSTRACT
TOP
 ABSTRACT
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
The central pathways and mediators involved in sympathoexcitatory responses to circulating leptin are not well understood, although the arcuate-paraventricular nucleus (ARC-PVN) pathway likely plays a critical role. In urethane-anesthetized rats, ipsilateral intracarotid artery (ICA) injection of murine leptin (100 µg/kg) activated most PVN neurons tested. These responses were reduced by intracerebroventricular injection of the melanocortin subtype 3 and 4 receptor (MC3/4-R) antagonist SHU-9119 (0.6 nmol). The MC3/4-R agonist MTII (0.6 nmol icv) activated PVN neurons. Some PVN neurons that were excited by ICA leptin were inhibited by local application of neuropeptide Y (NPY, 2.5 ng). ICA leptin (100 µg/kg) excited presympathetic rostral ventrolateral medulla neurons and renal sympathetic nerve activity without significant change in blood pressure or heart rate; these effects were mimicked by intracerebroventricular injection of MTII (0.6 nmol). These data provide in vivo electrophysiological evidence to support the hypothesis that circulating leptin activates the sympathetic nervous system by stimulating the release of {alpha}-melanocyte-stimulating hormone in the vicinity of PVN neurons that are inhibited by the orexogenic peptide NPY.

obesity; sympathetic nerve activity; extracellular recording; neuropeptide Y; {alpha}-melanocyte stimulating hormone

LEPTIN IS A PEPTIDE HORMONE that is produced by adipose tissue and acts centrally to decrease appetite and increase energy expenditure by activating the sympathetic nervous system. The central pathways mediating the sympathetic responses to leptin are still poorly understood. The paraventricular nucleus (PVN) of the hypothalamus, which has an important regulatory influence over sympathetic nerve activity, is thought to be a primary effector site for leptin's central actions (11, 14, 33). Among the proposed neuropeptide mediators of leptin's central actions are neuropeptide Y (NPY), which acts within the PVN to inhibit sympathetic nerve activity (9, 17, 21), and {alpha}-melanocyte-stimulating hormone ({alpha}-MSH), which acts within the forebrain to excite sympathetic nerve activity (18, 22). These two peptides, which are released into PVN by neurons originating in the arcuate nucleus (ARC), are likely to modulate the sympathoexcitatory responses of PVN neurons to blood-borne leptin. In the present study, we tested the hypothesis that leptin exerts its excitatory influence on sympathetic activity via release of {alpha}-MSH onto PVN neurons and that NPY acts on PVN neurons to reduce sympathetic activity. We recorded the responses of neurons in the PVN or in the rostral ventrolateral medulla (RVLM), an important medullary center for sympathetic drive, along with renal sympathetic nerve activity (RSNA) and arterial pressure (AP), to intra-arterial murine leptin injections targeting the forebrain region or to intracerebroventricular administration of a melanocortin receptor agonist and antagonist. Preliminary results of these studies have been reported in abstract form (34, 35).


   MATERIALS AND METHODS
TOP
 ABSTRACT
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
These studies were performed in accordance with the "Guiding Principles for Research Involving Animals and Human Beings" of the American Physiological Society (1). The experimental procedures were approved by the University of Iowa Institutional Animal Care and Use Committee.

All experiments were performed on adult male Sprague-Dawley rats (300-350 g; Harlan Sprague Dawley). The animals were housed in the University Animal Care Facility and exposed to a normal 12:12-h light-dark cycle.

General preparation. Rats were anesthetized with urethane (1.5 g/kg ip), and supplemental doses of urethane (0.1-0.3 g/kg ip or iv) were given when obvious increases in blood pressure and respiratory rate were observed during surgery or experimental recording. The level of anesthesia was periodically reassessed during the surgical procedures and experimental recording by examining nociceptive reflex responses and by continuously monitoring blood pressure and heart rate (HR). The left femoral artery was cannulated with PE-50 tubing filled with heparinized saline (20 U/ml) for the recording of arterial pressure, which was monitored with a Hewlett-Packard 7754A chart recorder (HP Medical Products Group, Andover, MA). The left femoral vein was cannulated with PE-20 tubing for the administration of drugs. A rostrally directed PE-20 cannula was inserted into the left common carotid artery, with the tip placed in the bifurcation for intracarotid artery (ICA) injection as previously reported (36, 37). This injection predominantly targets the ipsilateral forebrain (19) and therefore provides a means of delivering drugs to that region by the natural blood-borne route.

Animals were intubated and breathed spontaneously. Core temperature was maintained at 37 ± 0.2°C with a rectal thermometer and a temperature controller (model K-100, Baxter Healthcare, Valencia, CA). The head was fixed in a stereotaxic frame (David Kopf Instrument, Tujunga, CA).

Intracerebroventricular injection. In some animals, a 29-gauge stainless steel cannula was implanted into the left lateral cerebral ventricle [stereotaxic coordinates: 0.9-1.0 mm posterior to bregma; 1.4-1.6 mm lateral to midline; 3.2-3.3 mm ventral to dura (26)]. The intracerebroventricular position of the cannula was confirmed by the staining of all four ventricles after injection of 5 µl Pontamine sky blue at the end of the experiments.

Electrophysiological recording. A small craniotomy was made above the region of interest, and a glass micropipette was placed in the left PVN or left RVLM to record extracellular single-unit activity, using previously described techniques (36, 37). In one group of experiments, PVN recording electrode was attached to a second glass pipette (tip diameter ~50-100 µm) for regional microinjection of NPY into PVN. The tip of the microinjection pipette was placed approximately 0.2-0.4 mm proximal to the tip of the recording electrode. The microinjection pipette was connected to a Neurophore BH-2 (Medical Systems) pressure microinjection system. Volume of injection was measured at the time of delivery by observing the movement of a meniscus in the pipette through a horizontal microscope fitted with a calibrated eyepiece micrometer. Stereotaxic coordinates were: PVN, 1.6-2.1 mm posterior to bregma, 0.3-0.5 mm from midline, and 7.0-8.0 mm ventral to dura; RVLM, 11.8-12.8 mm posterior to bregma, 1.8-2.1 mm from midline, and 8.0-10.0 mm ventral to dura (26).

Recordings of RSNA were obtained from the left renal nerve using methods previously described from our lab (31, 37). In brief, the left kidney was exposed through a flank incision. A branch of the renal nerve was dissected free from surrounding tissue and placed on bipolar silver wire recording electrodes. When an optimal signal-to-noise ratio was established, the electrode and the renal nerve were covered with silicon sealant (World Precision Instruments, Sarasota, FL). The electrodes were sutured to the back muscles. Renal nerve activity was amplified (model P511, Grass Instrument; Quincy, MA) and displayed on an oscilloscope (TDS 3014, Tektronix, Beaverton, OR). The noise level was finally determined at the end of experiment after injection of hexamethonium (30 mg/kg iv). The net value of RSNA was calculated by subtracting the background noise from the actual recorded value during the experiment.

Experimental protocols. The recording session began at least 1 h after completion of the surgical preparation. PVN and RVLM neurons were tested initially for responses to blood pressure changes induced by an intravenous bolus of phenylephrine and nitroprusside, as previously described (36, 37). RVLM neurons were also tested for correlation with the peak arterial pressure pulse.

The first series of experiments was designed to determine the effects of leptin, and of the melanocortin receptors, as putative mediators of the leptin response, on PVN neurons. Murine leptin (100 µg/kg) was injected in the left ICA, and PVN neuronal activity and arterial pressure were recorded. In some of these neurons, the melanocortin antagonist SHU-9119 was administered intracerebroventricularly after the initial response to ICA leptin was observed. Other PVN neurons were tested for the effects of the melanocortin agonist MTII, administered intracerebroventricularly; some of these neurons were tested for the effect of intracerebroventricular administration of SHU-9119 after an initial response to MTII was observed. Some PVN neurons were also tested for their response to regional microinjection of NPY and to ICA leptin. In a second series of experiments, to determine the effect of leptin and MTII on downstream sympathetic output and cardiovascular variables, we recorded the responses of RVLM neuronal activity and simultaneously recorded RSNA, AP, and HR to ICA murine leptin and intracerebroventricular MTII.

The last unit recorded in each experiment was marked with iontophoresis of pontamine sky blue for subsequent determination of recording sites.

Data acquisition and analysis. The AP signal, the rectified and integrated voltage from the renal nerve recording, the transistor-transistor logic (TTL) pulses indicating multifiber action potentials in the raw RSNA exceeding a selected voltage, and the TTL pulses indicating PVN and RVLM single-unit activity were fed into an online data-acquisition system consisting of a Cambridge Electronics Design (CED, Cambridge, UK) 1401 Plus computer interface coupled with a Gateway Pentium personal computer. HR was derived from the peak AP tracing. Mean arterial pressure (MAP), HR, RSNA, and single-unit discharge were averaged over 1-min intervals. A 3-min baseline was used as control. Since the responses to local microinjection of NPY were of shorter duration, they were analyzed over a shorter time interval (30 s) using peak responses in PVN neuronal activity and MAP. Changes in RSNA were calculated as a percent change from the baseline activity. In the representative tracings, both integrated and windowed RSNA are shown, but only the integrated activity was used in the analysis of grouped data. A net percent change in PVN or RVLM neuronal discharge rate of ±30% or more was considered a significant effect of an experimental intervention as previously reported (36). Statistical significance among multiple comparisons was determined by one-way or two-way repeated-measures ANOVA followed by post hoc Fisher's least squares difference test. Student's t-test was employed for analysis of paired or unpaired data. Values are expressed as means ± SE. P < 0.05 was considered to indicate statistical significance.

Anatomy/histology. At the conclusion of each experiment, the rat was killed with an overdose of urethane. The brain was removed and fixed in a 10% formalin solution for at least 3 days and then sectioned (40 µm) on a cryostatic microtome (OM2563, Triangle Biomedical Sciences, Durham, NC). The sections were thaw-mounted on microscope slides and then stained with 1% aqueous neutral red. The last recording site marked with Pontamine sky blue was identified with a light microscope, and the locations of other recording sites were extrapolated with respect to this reference point. Recording sites were plotted on representative schematic tracings of the PVN and RVLM, based on the rat atlas of Paxinos and Watson (26).

Drugs. NPY, phenylephrine hydrochloride, SHU-9119, and sodium nitroprusside were purchased from Sigma (St. Louis, MO). Murine leptin was generously supplied by Amgen (Thousand Oaks, CA). MTII was purchased from Phoenix Pharmaceuticals (Belmont, CA). All drugs were dissolved in artificial cerebrospinal fluid (aCSF) for ICA or intracerebroventricular injection and in saline for intravenous injection. ICA injections were given in a volume of 10-20 µl flushed by 25 µl aCSF (pH 7.5). The volume for intracerebroventricular injection was 5-10 µl. The same volume of aCSF was administered via the ICA or intracerebroventricularly as a control.


   RESULTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 GRANTS
 REFERENCES
 
Effects of ICA murine leptin on PVN neuronal activity. Twenty-seven of 29 PVN neurons responded to ipsilateral ICA injection of murine leptin (100 µg/kg) with an increase in activity of at least 30% of baseline as previously reported (36). The onset latency of the response was 3-10 min, and the duration of the response was at least the 90 min tested. One representative PVN neuron is illustrated in Fig. 1A, showing gradual increase of neuronal firing after ICA leptin administration. The mean firing rate of neurons increased from baseline of 2.2 ± 0.3 to a peak of 4.0 ± 0.5 spikes/s 20 min after leptin injection (Fig. 1C, n = 27, P < 0.01). Two PVN neurons showed no change (<30%) in neuronal activity after ICA leptin. Mean arterial pressure (MAP) did not change significantly (Fig. 1C, from 97 ± 2 to 98 ± 2 mmHg) 20 min after leptin injection. The effects of ICA leptin were reproducible on a second injection. The same volume of aCSF injected via the ICA had no effects on PVN neuronal activity and MAP (n = 5, data not shown).



View larger version (32K):
[in this window]
[in a new window]
  		Fig. 1. A: representative paraventricular nucleus (PVN) neuron responding to ipsilateral intracarotid artery (ICA) injection of murine leptin. Leptin increases the PVN neuronal activity without significant change in arterial pressure (AP). B: representative tracing showing that the melanocortin subtype 3 and 4 receptor (MC3/4-R) antagonist SHU-9119 blocks the ongoing PVN neuronal response to ICA leptin injection. ICV, intracerebroventicular. C: grouped data showing the effect of MC3/4-R blockade on leptin-induced PVN neuronal response and mean AP (MAP) responses. *P < 0.05, vs. baseline at time 0. Values are means ± SE.

 

Effects of the MC3/4-R antagonist SHU-9119 on PVN neuronal responses to ICA murine leptin. Because {alpha}-MSH acting on MC4-R is a putative mechanism for the leptin-induced sympathetic response, we tested the effect of SHU-9119 on the PVN neuronal response to ICA leptin. Intracerebroventricular SHU-9119 (0.6 nmol) significantly reduced ongoing PVN neuronal responses to leptin (from 3.9 ± 0.6 to 2.2 ± 0.3 spikes/s, n = 12, P < 0.01, Fig. 1, B and C). Intracerebroventricular SHU-9119 alone had no effect on PVN neuronal activity and MAP (n = 3).

Effects of MTII on leptin-responsive PVN neurons. The melanocortin agonist MTII (0.6 nmol icv) increased PVN neuronal firing rate from a baseline of 2.0 ± 0.3 to a peak of 3.8 ± 0.6 spikes/s (n = 11, P < 0.01) within 10 min of injection (Fig. 2C) in 11 of 12 neurons tested. A typical example of PVN neuron responding to MTII is shown in Fig. 2A. The effect lasted ~30 min. Most of these neurons (7/9) also responded to ICA murine leptin. Blood pressure did not change after intracerebroventricular MTII injection (from 101 to 104 mmHg, n = 11). The MC3/4-R antagonist SHU-9119 (0.6 nmol icv) completely abolished the PVN neuronal response to MTII (from 3.8 ± 0.4 to 2.2 ± 0.3 spikes/s, n = 7, P < 0.01, Fig. 2, B and C). The effects of MTII were reproducible on a second intracerebroventricular injection.



View larger version (32K):
[in this window]
[in a new window]
  		Fig. 2. A: representative tracing showing that the MC3/4-R agonist (MTII) increases PVN neuronal activity with no significant change in AP. B: typical example showing that the MC3/4-R antagonist (SHU-9119) blocks the ongoing PVN response to MTII. C: grouped data showing the effects of the MC3/4-R antagonist on PVN neuronal and MAP responses to MTII. *P < 0.05, vs. baseline at time 0. Values are means ± SE.

 

Effects of local NPY on PVN neuronal activity. Local microinjection of NPY (2.5 ng in 50 nl) into the PVN from a pipette attached to the recording electrode decreased firing rate of 9 of 11 PVN neurons tested from 1.8 ± 0.3 to 0.9 ± 0.3 spikes/s (n = 9, P < 0.01) with only a slight fall in MAP (from 101 ± 4 to 98 ± 5 mmHg) as shown in Fig. 3. The duration of PVN neuronal responses to local NPY lasted only several minutes. Two PVN neurons did not respond to local microinjection of NPY. Six of the nine neurons inhibited by local NPY were excited by ICA murine leptin as shown in Fig. 3. The same volume of aCSF microinjections had no effects on PVN neuronal activity and MAP (n = 3).



View larger version (34K):
[in this window]
[in a new window]
  		Fig. 3. A: example of a PVN neuron inhibited by microinjection of neuropeptide Y (NPY, 2.5 ng, 50 nl) using attached second glass pipette (left). Slight decrease of AP occurred after microinjection of NPY within PVN region. This neuron also responded to ICA leptin by increase of PVN neuronal activity (right). B: summary of effects of microinjection of NPY (left; n = 9) within PVN area and ICA leptin (right; n = 6) on PVN neuronal activity and MAP. Leptin excited these NPY inhibited neurons. *P < 0.01. Values are means ± SE.

 

Effects of ICA murine leptin and intracerebroventricular MTII on simultaneously recorded RVLM neuronal activity, RSNA, AP, and HR. The above experiments suggest that leptin activates hypothalamic PVN neurons by releasing melanocortins in the PVN region. To determine whether these influences affect downstream neurons in sympathetic efferent pathways, we simultaneously recorded from RVLM neurons and the renal sympathetic outflow during ipsilateral ICA administration of murine leptin or intracerebroventricular injection of MTII.

Ipsilateral ICA administration of leptin (100 µg/kg) gradually increased RVLM neuronal activity and simultaneously increased RSNA, with only slight changes in AP and HR (Fig. 4). The peak responses occurred ~30 min after leptin injection. The RVLM neuronal firing rate increased from baseline of 10.3 ± 1.5 to a peak of 20.0 ± 2.2 spikes/s (P < 0.01, n = 12). Two RVLM neurons showed no changes in neuronal activity after leptin injection. Most of RVLM neurons tested had pulse-related firing pattern (Fig. 4B), suggesting that they were baroreceptor-sensitive presympathetic neurons. RSNA increased 25.4 ± 1.6% from baseline (P < 0.01, n = 12). The same volume of ICA injection of aCSF (n = 5, Fig. 4C) had no effects on RVLM firing rate, RSNA, AP, or HR.



View larger version (37K):
[in this window]
[in a new window]
  		Fig. 4. A: simultaneously recorded responses of AP, heart rate (HR), renal sympathetic nerve activity (RSNA, discharge rate and integrated voltage), and rostral ventrolateral medulla (RVLM) neuronal activity, showing the gradual increase in RVLM neuronal firing and RSNA after ICA administration of murine leptin without significant changes in AP and HR. bpm, Beats/min. B: cross correlations of RSNA and RVLM firing with peak AP, showing the pulse-related firing patterns. C: group data showing time course of changes in MAP, HR, RSNA and RVLM neuronal firing after ICA injection of leptin compared with same volume of artificial cerebrospinal fluid (aCSF) injection. Values are means ± SE.

 

Central administration of MTII (0.6 nmol icv, n = 7) activated RVLM neurons, increasing firing rate from a baseline of 13.6 ± 2.3 to a peak of 21.6 ± 2.0 spikes/s (P < 0.01) and increased RSNA integrated voltage by 31 ± 1.8% from baseline (P < 0.01) with an onset latency of ~2-3 min and a duration of effect of ~30 min (Fig. 5). There were no significant changes in AP and HR, although there is a trend toward elevation in MAP (Fig. 5B).



View larger version (23K):
[in this window]
[in a new window]
  		Fig. 5. A: simultaneous responses of RVLM neuronal activity, RSNA, AP, and HR showing the increase in RVLM neuronal firing and RSNA after intracerebroventricular administration of melanocortin agonist MTII, with slight elevations in HR. B: grouped data showing the time courses of the responses of MAP, HR, RSNA, and RVLM neuronal firing after intracerebroventricular injection of MTII compared with same volume of aCSF injection. Values are means ± SE.

 

Recording sites within PVN and RVLM regions. Figure 6 shows the locations of the single-unit recording sites in the PVN (Fig. 6A) and RVLM (Fig. 6B). The PVN neurons tested in this study were distributed mainly in the medial regions close to the third ventricle. There was no obvious clustering of neurons related to responses to administered drugs. Neurons recorded in the RVLM mostly were barosensitive and pulse related, although the specific function of these neurons are not known. Within the RVLM, there was no obvious difference in anatomic distribution between neurons activated by ICA murine leptin or intracerebroventricular MTII.



View larger version (33K):
[in this window]
[in a new window]
  		Fig. 6. Schematic reconstruction of recording sites within PVN (A) and RVLM (B) regions. Locations shown are leptin-responsive neurons ({bullet}) and nonresponsive neurons ({circ}) within PVN and RVLM; MTII-responsive PVN and RVLM neurons ({blacktriangleup}); PVN neurons responsive to both MTII and leptin ({blacksquare}); PVN neurons responsive to local NPY application ({diamondsuit}); PVN neurons responsive to both NPY and leptin (filled hexagon). The distance from bregma is indicated. Sections are modified from Paxinos and Watson (26). AH, anterior hypothalamus; f, fornix; IO, inferior olive; LPGi, lateral paragigantocellular nucleus; NA, nucleus ambiguus; NTS, nucleus of the solitary tract; py, pyramidal tract; 3V, third ventricle.

 


   DISCUSSION
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
GRANTS
 REFERENCES
 
The present study demonstrates that melanocortin receptors mediate the excitatory effects of circulating murine leptin on hypothalamic PVN neurons and that NPY has an opposing effect of inhibiting the activity of PVN neurons that are excited by blood-borne leptin. The study also demonstrates that acutely administered blood-borne leptin excites downstream brain stem presympathetic RVLM neurons and RSNA without inducing significant changes in blood pressure and HR and that these effects are mimicked by injecting the MC3/4-R agonist MTII into the cerebral ventricle. These results provide direct electrophysiological evidence in an in vivo model to support the hypothesis that blood-borne leptin activates the sympathetic nervous system by stimulating the release of {alpha}-MSH in the vicinity of PVN neurons that also respond to the suppressive effects of the orexogenic peptide NPY on sympathetic activity.

It is now generally agreed that leptin acts on the ARC to regulate the release of {alpha}-MSH and NPY into the PVN, the major hypothalamic effector site (3, 6) for leptin's actions. Leptin receptors have been identified on both NPY- and proopiomelanocortin (POMC)-expressing neurons in ARC (13, 28). Functional neuoranatomy studies using c-fos protein expression techniques have demonstrated that the PVN is the major hypothalamic region consistently activated by leptin regardless of dose, delivery route (intravenous or intracerebroventricular), and species (10-12). A recent electrophysiological study demonstrated that leptin increased the frequency of action potentials in POMC-expressing neurons through two mechanisms (7). Leptin causes a direct depolarization of POMC neurons through the activation of a nonspecific cation channel and simultaneously reduces the frequency of inhibitory postsynaptic current mediated by local NPY and GABAergic neurons. Thus leptin appears to have both pre- and postsynaptic actions that increase the neuronal activity of hypothalamic POMC neurons, which then increase endogenous melanocortin agonist release (e.g. {alpha}-MSH) within the PVN area. The critical role for the melanocortin system in leptin signaling emerged with the cloning of MC-4 receptor gene and the recognition that it is expressed primarily in the brain regions regulating autonomic functions, including the hypothalamic PVN (20, 25). In mice, intracerebroventricular administration of the synthetic MC3/4 receptor agonist MTII inhibits feeding, and this effect can be completely blocked by the melanocortin antagonist SHU-9119 (16). Furthermore, administration of SHU-9119 alone significantly enhanced feeding, indicating that central melanocortinergic neurons exert a tonic inhibition of feeding behavior (16).

The known connections of parvocellular PVN neurons with autonomic regions of the brain stem and spinal cord suggest that excitation of PVN neurons by leptin or melanocortin might result in increases in sympathetic nerve activity, AP, and HR. However, our data show that the intracerebroventricular melanocortin agonist MTII activated PVN or RVLM neurons and increased RSNA, but without significant changes in MAP and HR. This finding is consistent with that of Haynes et al. (18), who demonstrated that the acute administration of intracerebroventricular MTII increased sympathetic nerve traffic to brown adipose tissue and renal and lumbar beds without changing arterial pressure. In their study, SHU-9119 completely blocked leptin-induced renal but not brown adipose tissue sympathoexcitation, suggesting differential central pathways regulating leptin-induced sympathetic responses to different peripheral tissues. Microinjection of leptin into ventromedial hypothalamus (30) produced a similar result, with an increase in RSNA but no change in MAP. The explanation for this now well-recognized dissociation between MAP and RSNA in the response to acute administration of leptin is not known, but a possible explanation might be that the renal nerve subserves several physiological functions (8), all of which may not be linked to the immediate regulation of arterial pressure. In contrast, chronic intravenous or intracerebroventricular infusion of leptin can induce increases in blood pressure and heart rate in conscious rats (4, 5, 29).

Few prior studies have been done to test the central mechanism by which leptin excites SNA and the cardiovascular system. In our study, increased PVN neuronal activity induced by blood-borne murine leptin could be blocked by melanocortin receptor antagonist (SHU-9119), and RSNA could be stimulated by intracerebroventricular injection of MTII, strongly supporting the concept that the leptin signal driving renal sympathetic nerve activity is mediated by melanocortin receptors at the hypothalamic level. In this study, we did not measure sympathetic drive to brown adipose tissue, which has a primarily thermogenic/metabolic function. Leptin activation of that sympathetic pathway may utilize an alternate central pathway via raphe pallidus (23, 24).

The mechanism by which released NPY inhibits PVN neurons is not well known. In the present study, regional application of NPY within PVN significantly reduced the PVN neuronal activity, and more specifically inhibited the activity of PVN neurons that also responded to ICA leptin. A recent study in hypothalamus suggested that NPY induces the release of GABA, with GABA-mediated presynaptic inhibition (6). Inhibition might also be mediated postsynaptically via activation of local inhibitory interneurons or via a direct influence of NPY on the recorded neurons. These possibilities were not tested in this study. In leptin-deficient mice, the NPY levels in the hypothalamus are dramatically increased (32), while increasing leptin inhibits NPY gene expression (15). This reciprocal relationship between leptin and NPY further supports the role of NPY as an inhibitory mediator of leptin-related functions. Leptin and NPY have opposite effects on many other autonomic functions (2, 27). Interactions between leptin and NPY have been found in the control of RSNA and blood pressure (21), but in the present study this relationship was not directly examined.

In summary, we have demonstrated that the sympathoexcitatory response to blood-borne leptin, delivered acutely via the intracarotid route, is mediated by melanocortin receptors in the hypothalamic paraventricular region. We have also demonstrated that systemically administered murine leptin can excite PVN neurons that are inhibited by NPY. These results provide strong electrophysiological support for the general hypothesis that {alpha}-MSH and NPY interact in a reciprocal fashion within the PVN to modulate the sympathetic response to leptin.


   GRANTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
REFERENCES
 
This work was supported by NIH Program Project Grant PO1 HL-014388 (Principal Investigator, F. M. Abboud; Project Director, R. B. Felder).


   ACKNOWLEDGMENTS
 
Murine leptin was supplied through an agreement with Amgen.


   FOOTNOTES
 

Address for reprint requests and other correspondence: R. B. Felder, Univ. of Iowa College of Medicine, E318-GH, 200 Hawkins Dr., Iowa City, IA 52242 (E-mail: robert-felder{at}uiowa.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.


   REFERENCES
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 

  1. American Physiological Society. Guiding principles for research involving animals and human beings. Am J Physiol Regul Integr Comp Physiol 283: R281-R283, 2002.[Free Full Text]
  2. Ammar AA, Sederholm F, Saito TR, Scheurink AJ, Johnson AE, and Sodersten P. NPY-leptin: opposing effects on appetitive and consummatory ingestive behavior and sexual behavior. Am J Physiol Regul Integr Comp Physiol 278: R1627-R1633, 2000.[Abstract/Free Full Text]
  3. Broberger C, Johansen J, Johansson C, Schalling M, and Hokfelt T. The neuropeptide Y/agouti gene-related protein (AGRP) brain circuitry in normal, anorectic, and monosodium glutamate-treated mice. Proc Natl Acad Sci USA 95: 15043-15048, 1998.[Abstract/Free Full Text]
  4. Carlyle M, Jones OB, Kuo JJ, and Hall JE. Chronic cardiovascular and renal actions of leptin: role of adrenergic activity. Hypertension 39: 496-501, 2002.[Abstract/Free Full Text]
  5. Correia ML, Morgan DA, Sivitz WI, Mark AL, and Haynes WG. Leptin acts in the central nervous system to produce dose-dependent changes in arterial pressure. Hypertension 37: 936-942, 2001.[Abstract/Free Full Text]
  6. Cowley MA, Pronchuk N, Fan W, Dinulescu DM, Colmers WF, and Cone RD. Integration of NPY, AGRP, and melanocortin signals in the hypothalamic paraventricular nucleus: evidence of a cellular basis for the adipostat. Neuron 24: 155-163, 1999.[CrossRef][Web of Science][Medline]
  7. Cowley MA, Smart JL, Rubinstein M, Cerdan MG, Diano S, Horvath TL, Cone RD, and Low MJ. Leptin activates anorexigenic POMC neurons through a neural network in the arcuate nucleus. Nature 411: 480-484, 2001.[CrossRef][Medline]
  8. DiBona GF. Central angiotensin modulation of baroreflex control of renal sympathetic nerve activity in the rat: influence of dietary sodium. Acta Physiol Scand 177: 285-289, 2003.[CrossRef][Web of Science][Medline]
  9. Egawa M, Yoshimatsu H, and Bray GA. Neuropeptide Y suppresses sympathetic activity to interscapular brown adipose tissue in rats. Am J Physiol Regul Integr Comp Physiol 260: R328-R334, 1991.[Abstract/Free Full Text]
  10. Elias CF, Kelly JF, Lee CE, Ahima RS, Drucker DJ, Saper CB, and Elmquist JK. Chemical characterization of leptin-activated neurons in the rat brain. J Comp Neurol 423: 261-281, 2000.[CrossRef][Web of Science][Medline]
  11. Elmquist JK. Hypothalamic pathways underlying the endocrine, autonomic, and behavioral effects of leptin. Physiol Behav 74: 703-708, 2001.[CrossRef][Medline]
  12. Elmquist JK, Ahima RS, Maratos-Flier E, Flier JS, and Saper CB. Leptin activates neurons in ventrobasal hypothalamus and brainstem. Endocrinology 138: 839-842, 1997.[Abstract/Free Full Text]
  13. Elmquist JK, Bjorbaek C, Ahima RS, Flier JS, and Saper CB. Distributions of leptin receptor mRNA isoforms in the rat brain. J Comp Neurol 395: 535-547, 1998.[CrossRef][Web of Science][Medline]
  14. Elmquist JK, Maratos-Flier E, Saper CB, and Flier JS. Unraveling the central nervous system pathways underlying responses to leptin. Nat Neurosci 1: 445-450, 1998.[CrossRef][Web of Science][Medline]
  15. Erickson JC, Clegg KE, and Palmiter RD. Sensitivity to leptin and susceptibility to seizures of mice lacking neuropeptide Y. Nature 381: 415-421, 1996.[CrossRef][Medline]
  16. Fan W, Boston BA, Kesterson RA, Hruby VJ, and Cone RD. Role of melanocortinergic neurons in feeding and the agouti obesity syndrome. Nature 385: 165-168, 1997.[CrossRef][Medline]
  17. Harland D, Bennett T, and Gardiner SM. Cardiovascular actions of neuropeptide Y in the hypothalamic paraventricular nucleus of conscious Long Evans and Brattleboro rats. Neurosci Lett 85: 239-243, 1988.[CrossRef][Web of Science][Medline]
  18. Haynes WG, Morgan DA, Djalali A, Sivitz WI, and Mark AL. Interactions between the melanocortin system and leptin in control of sympathetic nerve traffic. Hypertension 33: 542-547, 1999.[Abstract/Free Full Text]
  19. Haywood JR, Fink GD, Buggy J, Phillips MI, and Brody MJ. The area postrema plays no role in the pressor action of angiotensin in the rat. Am J Physiol Heart Circ Physiol 239: H108-H113, 1980.[Abstract/Free Full Text]
  20. Kishi T, Aschkenasi CJ, Lee CE, Mountjoy KG, Saper CB, and Elmquist JK. Expression of melanocortin 4 receptor mRNA in the central nervous system of the rat. J Comp Neurol 457: 213-235, 2003.[CrossRef][Web of Science][Medline]
  21. Matsumura K, Tsuchihashi T, and Abe I. Central cardiovascular action of neuropeptide Y in conscious rabbits. Hypertension 36: 1040-1044, 2000.[Abstract/Free Full Text]
  22. Matsumura K, Tsuchihashi T, Abe I, and Iida M. Central {alpha}-melanocyte-stimulating hormone acts at melanocortin-4 receptor to activate sympathetic nervous system in conscious rabbits. Brain Res 948: 145-148, 2002.[CrossRef][Web of Science][Medline]
  23. Morrison SF. Differential regulation of brown adipose and splanchnic sympathetic outflows in rat: roles of raphe and rostral ventrolateral medulla neurons. Clin Exp Pharmacol Physiol 28: 138-143, 2001.[CrossRef][Web of Science][Medline]
  24. Morrison SF. Differential regulation of sympathetic outflows to vasoconstrictor and thermoregulatory effectors. Ann NY Acad Sci 940: 286-298, 2001.[Web of Science][Medline]
  25. Mountjoy KG, Mortrud MT, Low MJ, Simerly RB, and Cone RD. Localization of the melanocortin-4 receptor (MC4-R) in neuroendocrine and autonomic control circuits in the brain. Mol Endocrinol 8: 1298-1308, 1994.[Abstract/Free Full Text]
  26. Paxinos G and Watson C. The Rat Brain in Stereotaxic Coordinates. San Diego, CA: Academic, 1986.
  27. Schwartz GJ and Moran TH. Leptin and neuropeptide Y have opposing modulatory effects on nucleus of the solitary tract neurophysiological responses to gastric loads: implications for the control of food intake. Endocrinology 143: 3779-3784, 2002.[Abstract/Free Full Text]
  28. Schwartz MW, Seeley RJ, Campfield LA, Burn P, and Baskin DG. Identification of targets of leptin action in rat hypothalamus. J Clin Invest 98: 1101-1106, 1996.[Web of Science][Medline]
  29. Shek EW, Brands MW, and Hall JE. Chronic leptin infusion increases arterial pressure. Hypertension 31: 409-414, 1998.[Abstract/Free Full Text]
  30. Tanida M, Nagai K, and Kaneko H. Activation of the renal sympathetic nerve by leptin microinjection into the ventromedial hypothalamus in rats. In Vivo 17: 213-217, 2003.[Web of Science][Medline]
  31. Wei SG and Felder RB. Forebrain renin-angiotensin system has a tonic excitatory influence on renal sympathetic nerve activity. Am J Physiol Heart Circ Physiol 282: H890-H895, 2002.[Abstract/Free Full Text]
  32. Wilding JP, Gilbey SG, Bailey CJ, Batt RA, Williams G, Ghatei MA, and Bloom SR. Increased neuropeptide-Y messenger ribonucleic acid (mRNA) and decreased neurotensin mRNA in the hypothalamus of the obese (ob/ob) mouse. Endocrinology 132: 1939-1944, 1993.[Abstract/Free Full Text]
  33. Woods AJ and Stock MJ. Leptin activation in hypothalamus. Nature 381: 745, 1996.[Medline]
  34. Zhang ZH and Felder RB. Leptin activates hypothalamic paraventricular nucleus (PVN) neurons via the brain melanocortin system (Abstract). Soc Neurosci Abstr 27: 733, 2001.
  35. Zhang ZH and Felder RB. Opposite effects of leptin and NPY on hypothalamic paraventricular nucleus (PVN) neurons (Abstract). FASEB J 15: A77, 2001.
  36. Zhang ZH, Francis J, Weiss RM, and Felder RB. The renin-angiotensin-aldosterone system excites hypothalamic paraventricular nucleus neurons in heart failure. Am J Physiol Heart Circ Physiol 283: H423-H433, 2002.[Abstract/Free Full Text]
  37. Zhang ZH, Wei SG, Francis J, and Felder RB. Cardiovascular and renal sympathetic activation by blood-borne TNF-{alpha} in rat: the role of central prostaglandins. Am J Physiol Regul Integr Comp Physiol 284: R916-R927, 2003.[Abstract/Free Full Text]



This article has been cited by other articles:


Home page
 Am. J. Physiol. Heart Circ. Physiol.Home page
Z.-H. Zhang, Y. Yu, Y.-M. Kang, S.-G. Wei, and R. B. Felder
Aldosterone acts centrally to increase brain renin-angiotensin system activity and oxidative stress in normal rats
Am J Physiol Heart Circ Physiol, February 1, 2008; 294(2): H1067 - H1074.
[Abstract] [Full Text] [PDF]

Home page
 HypertensionHome page
S. D. Stocker, R. Meador, and J. M. Adams
Neurons of the Rostral Ventrolateral Medulla Contribute to Obesity-Induced Hypertension in Rats
Hypertension, March 1, 2007; 49(3): 640 - 646.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Web of Science (11)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Zhang, Z.-H.
Right arrow Articles by Felder, R. B.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Zhang, Z.-H.
Right arrow Articles by Felder, R. B.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Visit Other APS Journals Online
Copyright © 2004 by the American Physiological Society.