Feeding behavior elicited by mu opioid and GABA receptor activation in the lateral septum.

The lateral septum (LS), a brain region typically associated with behaviors involving reward, anxiety-like behavior, learning, and memory, has recently received increased interest due to its potential role in eating behavior. Our current results showed that morphine (5 µg) microinjected into the LS produced a stable feeding response. Specifically, across five days of repeated injections, there was no increase or sensitization effect, nor a decrease in feeding or tolerance. Additionally, we found that pretreatment with the broad-spectrum opioid receptor antagonist naloxone blocked morphine-elicited feeding, further supporting a role for LS opioid receptors in the activation of feeding behaviors. We had previously found that the GABAA receptor agonist muscimol produces a similar increase in feeding when injected into the LS. Given the involvement of the LS in multiple behaviors, we next evaluated whether other behaviors might be co-occurring with feeding in response to opioid or GABAA receptor agonist injection into the LS. We assessed eating, drinking, grooming, sleeping, activity levels and resting behavior for 3 h after injection of aCSF, DAMGO, morphine, or muscimol. We found that morphine and muscimol both decreased the latency to eat, and all drugs tested increased food intake. The feeding occurred within 30 min of muscimol injection but was delayed after opioid injections. The absence of increases in other goal-oriented behavior like drinking or grooming or behavioral hyperactivity supports a primary effect of muscimol and the opioids on LS mechanisms of feeding control. SIGNIFICANCE STATEMENT: The LS is interesting because of its role in a wide range of behaviors including defensive behaviors, social behaviors, learning, memory, and motivation. Although the LS was discovered to have a role in feeding stimulation over 30 years ago, only recently has major progress begun to reveal the underlying mechanisms. The present paper contributes by suggesting that LS GABAA and µ-opioid receptors elicit eating by inhibiting LS neurons that themselves inhibit eating. Importantly, this work informs lateral septal research which may shed light on disordered eating included binge eating and anorexia.

GLT-1-Dependent Disruption of CNS Glutamate Homeostasis and Neuronal Function by the Protozoan Parasite Toxoplasma gondii.

The immune privileged nature of the CNS can make it vulnerable to chronic and latent infections. Little is known about the effects of lifelong brain infections, and thus inflammation, on the neurological health of the host. Toxoplasma gondii is a parasite that can infect any mammalian nucleated cell with average worldwide seroprevalence rates of 30%. Infection by Toxoplasma is characterized by the lifelong presence of parasitic cysts within neurons in the brain, requiring a competent immune system to prevent parasite reactivation and encephalitis. In the immunocompetent individual, Toxoplasma infection is largely asymptomatic, however many recent studies suggest a strong correlation with certain neurodegenerative and psychiatric disorders. Here, we demonstrate a significant reduction in the primary astrocytic glutamate transporter, GLT-1, following infection with Toxoplasma. Using microdialysis of the murine frontal cortex over the course of infection, a significant increase in extracellular concentrations of glutamate is observed. Consistent with glutamate dysregulation, analysis of neurons reveal changes in morphology including a reduction in dendritic spines, VGlut1 and NeuN immunoreactivity. Furthermore, behavioral testing and EEG recordings point to significant changes in neuronal output. Finally, these changes in neuronal connectivity are dependent on infection-induced downregulation of GLT-1 as treatment with the ß-lactam antibiotic ceftriaxone, rescues extracellular glutamate concentrations, neuronal pathology and function. Altogether, these data demonstrate that following an infection with T. gondii, the delicate regulation of glutamate by astrocytes is disrupted and accounts for a range of deficits observed in chronic infection.

Direct hypothalamic and indirect trans-pallidal, trans-thalamic, or trans-septal control of accumbens signaling and their roles in food intake.

Due in part to the increasing incidence of obesity in developed nations, recent research aims to elucidate neural circuits that motivate humans to overeat. Earlier research has described how the nucleus accumbens shell (AcbSh) motivates organisms to feed by activating neuronal populations in the lateral hypothalamus (LH). However, more recent research suggests that the LH may in turn communicate with the AcbSh, both directly and indirectly, to re-tune the motivation to consume foods with homeostatic and food-related sensory signals. Here, we discuss the functional and anatomical evidence for an LH to AcbSh connection and its role in eating behaviors. The LH appears to modulate Acb activity directly, using neurotransmitters such as hypocretin/orexin or melanin concentrating hormone (MCH). The LH also indirectly regulates AcbSh activity through certain subcortical "relay" regions, such as the lateral septum (LS), ventral pallidum (VP), and paraventricular thalamus, using a variety of neurotransmitters. This review aims to summarize studies on these topics and outline a model by which LH circuits processing energy balance can modulate AcbSh neural activity to regulate feeding behavior.

Behaviorally specific versus non-specific suppression of accumbens shell-mediated feeding by ipsilateral versus bilateral inhibition of the lateral hypothalamus.

The nucleus accumbens shell (AcbSh) and lateral hypothalamus (LH) are linked in the control of food intake. Pharmacological inhibition of the LH may block AcbSh-elicited feeding, but the behavioral phenotype associated with this feeding suppression is unknown. To examine this phenotype, adult male Sprague-Dawley rats were implanted with three cannulas - one unilaterally in the AcbSh and two bilaterally in the LH - to allow for central drug injections. The AcbSh received injections of the AMPA receptor antagonist DNQX or the GABAA receptor agonist muscimol, while the LH received injections of the NMDA receptor antagonist D-AP5 or muscimol. Eating, drinking, grooming, locomotion, quiescence, and sleeping behaviors were measured every minute for 60 min post-injection. From these observational data, feeding bout durations, feeding frequency, and latency to feed were determined. AcbSh muscimol or DNQX increased food intake by increasing feeding bout durations and frequency and decreasing latency to feed. D-AP5 or muscimol, injected into the LH bilaterally or ipsilateral to the AcbSh injection, reversed these AcbSh-mediated effects. Though bilateral LH D-AP5 or muscimol injections blocked feeding responses, they also hastened onset of sleep. In contrast, ipsilateral LH D-AP5 or muscimol injections suppressed AcbSh-mediated feeding behaviors without substantially altering sleeping or other behaviors. These results suggest bilateral LH inhibition via NMDA receptor blockade or GABAA receptor activation produces behavioral effects that might indirectly suppress feeding, but ipsilateral LH inhibition through these receptors suppresses AcbSh AMPA and GABAA receptor-mediated feeding specifically. This evidence strengthens the concept of a feeding-specific association between these regions.

Ipsilateral feeding-specific circuits between the nucleus accumbens shell and the lateral hypothalamus: regulation by glutamate and GABA receptor subtypes.

The nucleus accumbens shell (AcbSh) and the lateral hypothalamus (LH) are both involved in the control of food intake. Activation of GABA(A) receptors or blockade of AMPA and kainate receptors within the AcbSh induces feeding, as does blockade of GABA(A) receptors or activation of NMDA receptors in the LH. Further, evidence suggests that feeding induced via the AcbSh can be suppressed by LH inhibition. However, it is unclear if this suppression is specific to feeding. Adult male Sprague-Dawley rats with 3 intracranial guide cannulas, one unilaterally into the AcbSh and two bilaterally into the LH, were used to explore this issue. DNQX (1.25 µg) or muscimol (100 ng) infused into the AcbSh unilaterally elicited feeding, and this elicited intake was suppressed by bilateral LH injection of d-AP5 (2 µg) or muscimol (25 ng). The effectiveness of d-AP5 or muscimol infusion into either the LH site ipsilateral or contralateral to the AcbSh injection was compared. Ipsilateral LH injection of d-AP5 or muscimol was significantly more effective than contralateral injection in suppressing food intake initiated by AcbSh injection of DNQX or muscimol. These results add to the prior evidence that inhibition of the LH through pharmacological modulation of NMDA or GABA(A) receptors specifically suppresses feeding initiated by AcbSh inhibition, and that these two regions communicate via an ipsilateral circuit to specifically regulate feeding.

Stimulation of lateral hypothalamic AMPA receptors may induce feeding in rats.

Glutamate or its ionotropic receptor (iGluR) agonists, N-methyl-D-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxale propionate (AMPA), and kainate (KA) elicit feeding when microinjected into the lateral hypothalamus (LH) of satiated rats. In the present study we investigated the contributions of AMPA and KA receptors (AMPARs and KARs) to feeding initiation. Intense feeding was elicited by LH injection of RS-AMPA (1 and 10 nmol) but not by the isolated, inactive R-AMPA enantiomer (1 and 10 nmol). Further, LH pretreatment with either the non-selective AMPAR/KAR antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 4 nmol) or the selective AMPAR antagonist, GYKI 52466 (10 nmol), suppressed AMPA-elicited food intake and, when combined, blocked AMPA-elicited food intake. These findings suggest that LH AMPARs mediate AMPA injection-elicited feeding with a possible contribution by KARs. In contrast, CNQX or GYKI 52466 injected into the LH at the onset of the nocturnal period or into fasted rats did not suppress the feeding produced by either condition. RS-AMPA injected into the LH of fasted or nocturnal feeding subjects elicited eating in both conditions; however, the magnitude of the increase was greater in fasted rats. These data suggest that selective stimulation of AMPAR in the LH is sufficient to elicit feeding. In contrast, the results did not provide evidence that AMPAR stimulation is necessary for deprivation-induced or nocturnal eating; however, they did suggest that modulatory interactions may exist between these receptors and these forms of naturally occurring eating behavior.

The tuberal lateral hypothalamus is a major target for GABAA--but not GABAB-mediated control of food intake.

The lateral hypothalamus (LH) is a site of integration for control mechanisms of feeding behavior as it has extensive reciprocal connections with multiple intrahypothalamic and extrahypothalamic brain areas. Evidence suggests that blockade of ionotropric gamma-aminobutyric acid (GABA) receptors in the LH elicits eating in satiated rats. To determine whether this GABA(A) receptor antagonist effect is specific to the LH, the antagonist picrotoxin was injected into one of six nearby sites and food intake was measured. Picrotoxin at 133 pmol elicited eating in the LH, but not in surrounding sites (thalamus, lateral preoptic area, ventral tegmental area, dorsomedial hypothalamus, and entopeduncular nucleus). More specifically, picrotoxin injected into the tuberal LH (tLH) elicited eating, but was ineffective when injected into the anterior or posterior LH. We also investigated whether GABA(B) receptors in the LH participated in the control of food intake and found that neither blockade nor activation of these receptors under multiple conditions changed food intake. Collectively, our findings suggest that GABA(A) but not GABA(B) receptors in the tLH act to suppress feeding behavior.

GABA(A) receptors in the lateral hypothalamus as mediators of satiety and body weight regulation.

In the lateral hypothalamus (LH), the inhibitory amino acid neurotransmitter, GABA, has had a long-standing presumptive role as an inhibitor of food intake. However, minimal investigation has been focused on GABA, especially as compared to the attention received by many peptide transmitters. To begin to address this deficiency in the understanding of the role of GABA in the LH and feeding, we report that antagonism of GABA(A) receptors in the rat LH elicits feeding, consistent with previous findings, and provide evidence for the behavioral selectivity of this effect. We extend previous findings that activation of LH GABA(A) receptors suppresses feeding, in particular by showing that nighttime and deprivation-induced eating are dramatically suppressed. Finally, we show that chronic activation, but not blockade, of the LH GABA(A) receptors leads to a reduction in 24 h food intake with concomitant body weight loss. These data collectively suggest that activation of GABA(A) receptors plays a fundamental role in controlling food intake and body weight, a role that has previously been somewhat underestimated.

Stimulation of lateral hypothalamic kainate receptors selectively elicits feeding behavior.

Glutamate and its receptor agonists, NMDA, AMPA, and KA, elicit feeding when microinjected into the lateral hypothalamus (LH) of satiated rats. However, determining the relative contributions of AMPA receptors (AMPARs) and KA receptors (KARs) to LH feeding mechanisms has been difficult due to a lack of receptor selective agonists and antagonists. Furthermore, LH injection of KA produces behavioral hyperactivity, questioning a role for KARs in feeding selective stimulation. In the present study, we used the KAR agonist, (RS)-2-amino-3-(3-hydroxy-5-tert-butylisoxazol-4-yl) propanoic acid (ATPA), which selectively binds the GluR5 subunit of KARs, to stimulate feeding, presumably via KAR activation. Using ATPA, we tested whether: (1) LH injection of ATPA elicits feeding, (2) prior treatment with the non-selective AMPA/KAR antagonist, CNQX, suppresses ATPA-elicited feeding, and (3) LH injection of ATPA elicits behavioral patterns specific for feeding. We found that injection of ATPA (0.1 and 1 nmol) elicited an intense feeding response (e.g., 4.8+/-1.6 g) that was blocked by LH pretreatment with CNQX, but was unaffected by pretreatment with the AMPAR selective antagonist, GYKI 52466. Furthermore, minute-by-minute behavioral analysis revealed that LH injection of ATPA increased time spent feeding to 55% of the initial test period with little or no effects on other behaviors at any time. In contrast, LH injection of KA similarly increased feeding but also produced intense locomotor activity. These data suggest that selective activation of LH KARs containing GluR5 subunit(s) is sufficient to elicit feeding.

Catecholaminergic control of mitogen-activated protein kinase signaling in paraventricular neuroendocrine neurons in vivo and in vitro: a proposed role during glycemic challenges.

Paraventricular hypothalamic (PVH) corticotropin-releasing hormone (CRH) neuroendocrine neurons mount neurosecretory and transcriptional responses to glycemic challenges [intravenous 2-deoxyglucose (2-DG) or insulin]. Although these responses require signals from intact afferents originating from hindbrain CA (catecholaminergic) neurons, the identity of these signals and the mechanisms by which they are transduced by PVH neurons during glycemic challenge remain unclear. Here, we tested whether the prototypical catecholamine, norepinephrine (NE), can reproduce PVH neuroendocrine responses to glycemic challenge. Because these responses include phosphorylation of p44/42 mitogen-activated protein (MAP) kinases [extracellular signal-regulated kinases 1/2 (ERK1/2)], we also determined whether NE activates ERK1/2 in PVH neurons and, if so, by what mechanism. We show that systemic insulin and 2-DG, and PVH-targeted NE microinjections, rapidly elevated PVH phospho-ERK1/2 levels. NE increased Crh and c-fos expression, together with circulating ACTH/corticosterone. However, because injections also increased c-Fos mRNA in other brain regions, we used hypothalamic slices maintained in vitro to clarify whether NE activates PVH neurons without contribution of inputs from distal regions. In slices, bath-applied NE triggered robust phospho-ERK1/2 immunoreactivity in PVH (including CRH) neurons, which attenuated markedly in the presence of the alpha1 adrenoceptor antagonist, prazosin, or the MAP kinase kinase (MEK) inhibitor, U0126 (1,4-diamino-2,3-dicyano-1,4-bis[2-aminophenylthio]butadiene). Therefore, at a systems level, local PVH delivery of NE is sufficient to account for hindbrain activation of CRH neuroendocrine neurons during glycemic challenge. At a cellular level, these data provide the first demonstration that MAP kinase signaling cascades (MEK-->ERK) are intracellular transducers of noradrenergic signals in CRH neurons, and implicate this transduction mechanism as an important component of central neuroendocrine responses during glycemic challenge.

NMDA receptors mediate feeding elicited by neuropeptide Y in the lateral and perifornical hypothalamus.

Neuropeptide Y (NPY) and N-methyl-d-aspartate (NMDA) receptors in the lateral (LH) and perifornical hypothalamus (PFH) are believed to be involved in the stimulation of feeding behavior. To investigate the possibility that neurons with these receptors interact to stimulate eating, the NMDA receptor antagonists d-(-)-2-amino-5-phosphonopentanoic acid (D-AP5) or 7-chlorokynurenic acid (7-CK) were injected into the LH or PFH of satiated rats 5 min prior to NPY in the same site and subsequent food intake was measured 1, 2, and 4 h postinjection. The injection of NPY (78 pmol/0.3 microl aCSF) in the PFH produced an average food intake of 9.7 g in 4 h, compared to the intake of 1.3 g after the artificial cerebrospinal fluid (aCSF) vehicle. D-AP5 (1, 10, or 20 nmol/0.3 microl aCSF) pretreatment suppressed NPY-induced eating, with the 20 nmol dose of D-AP5 producing up to an 80% suppression of elicited food intake down to 1.9 g in 4 h. Similar effects were produced with the LH as the injection site. Illustrating the specificity of the NMDA receptor antagonist's suppression of NPY-elicited feeding, D-AP5 suppressed NMDA-elicited feeding but did not affect the eating response induced by kainic acid. Consistent with the effects of D-AP5, the NMDA receptor antagonist 7-CK (40 nmol/0.3 microl dimethyl sulfoxide, DMSO) suppressed feeding elicited by NPY in the LH by 78%. Collectively, the findings suggest that the feeding elicited by NPY is dependent upon the activation of the NMDA receptors in the LH and PFH.

Origins of lateral hypothalamic afferents associated with N-methyl-d-aspartic acid-elicited eating studied using reverse microdialysis of NMDA and Fluorogold.

Afferent projections to the tuberal lateral hypothalamus (tLH), where excitatory amino acid application is most effective in eliciting feeding, and to the anterior, posterior and medial regions of the hypothalamus were studied using reverse microdialysis of N-methyl-D-aspartic acid (NMDA) and Fluorogold (FG). NMDA at 660 microM delivered for 10 min was effective in stimulating food intake only when administered into the tLH, causing a mean intake of 9.3 g compared to less than 1 g in any other site. Subsequent administration of FG through the dialysis probe retrogradely in labeled neurons in brain structures associated with the feeding response including the frontal cortex, amygdala, nucleus accumbens (NA), preoptic areas, substantia nigra, ventral tegmental area (VTA), parabrachial nucleus, and the nucleus of the solitary tract (NST). Labeling after anterior and posterior LH infusion of FG was similar to that seen after tLH delivery with some apparent differences, whereas FG administration into the medial hypothalamus produced a distinctly different pattern of labeling compared to the other groups. Some of the observed labeling appeared to be almost exclusively associated with the tLH where NMDA elicits feeding. In particular, amygdala, preoptic area and shell of the accumbens labeling was noticeably denser in tLH eaters than in all other groups. These findings are consistent with the role of LH glutamate and NMDA receptors in the regulation of food intake and identify afferents to the region which possibly mediate endogenous LH glutamate's effects on feeding.

The NMDA receptor antagonist MK-801 alters lipoprivic eating elicited by 2-mercaptoacetate.

Eating behavior is controlled, at least in part, by levels of circulating metabolic fuels such as glucose and free fatty acids, and drugs that interfere with the availability of these fuels can elicit eating. One such drug is 2-mercaptoacetate (2MA), an inhibitor of fatty acid oxidation. Evidence also suggests that NMDA receptors may mediate some aspects of normal eating and satiety. The present study was conducted in order to determine whether NMDA receptors may play a role in feeding elicited by 2MA. Rats received intraperitoneal injections of either saline, 2MA, the non-competitive NMDA receptor antagonist MK-801 or a combined injection of 2MA and MK-801, and subsequent intake of a fat-enriched, mash diet was measured at 1, 2, 3 and 4 h post-injection. Results showed that cumulative food intake was significantly increased by 2MA alone, as compared to saline controls, with most of the 2MA-elicited eating occurring during the first hour post-injection. While MK-801 alone did not alter food intake, it did have a biphasic effect on feeding elicited by 2MA. MK-801 initially suppressed and later enhanced eating elicited by 2MA. Although it is unclear whether MK-801 is acting centrally, peripherally or both to alter 2MA-induced eating, these results implicate NMDA receptors and the neurotransmitter glutamate in the regulation of lipid-associated eating and satiety.

Lateral hypothalamic signaling mechanisms underlying feeding stimulation: differential contributions of Src family tyrosine kinases to feeding triggered either by NMDA injection or by food deprivation.

In rats, feeding can be triggered experimentally using many approaches. Included among these are (1) food deprivation and (2) acute microinjection of the neurotransmitter l-glutamate (Glu) or its receptor agonist NMDA into the lateral hypothalamic area (LHA). Under both paradigms, the NMDA receptor (NMDA-R) within the LHA appears critically involved in transferring signals encoded by Glu to stimulate feeding. However, the intracellular mechanisms underlying this signal transfer are unknown. Because protein-tyrosine kinases (PTKs) participate in NMDA-R signaling mechanisms, we determined PTK involvement in LHA mechanisms underlying both types of feeding stimulation through food intake and biochemical measurements. LHA injections of PTK inhibitors significantly suppressed feeding elicited by LHA NMDA injection (up to 69%) but only mildly suppressed deprivation feeding (24%), suggesting that PTKs may be less critical for signals underlying this feeding behavior. Conversely, food deprivation but not NMDA injection produced marked increases in apparent activity for Src PTKs and in the expression of Pyk2, an Src-activating PTK. When considered together, the behavioral and biochemical results demonstrate that, although it is easier to suppress NMDA-elicited feeding by PTK inhibitors, food deprivation readily drives PTK activity in vivo. The latter result may reflect greater PTK recruitment by neurotransmitter receptors, distinct from the NMDA-R, that are activated during deprivation-elicited but not NMDA-elicited feeding. These results also demonstrate how the use of only one feeding stimulation paradigm may fail to reveal the true contributions of signaling molecules to pathways underlying feeding behavior in vivo.

Dual roles in feeding for AMPA/kainate receptors: receptor activation or inactivation within distinct hypothalamic regions elicits feeding behavior.

We have previously shown that hypothalamic injections of glutamate, or agonists of its ionotropic receptors (iGluRs), elicit intense feeding responses in satiated rats [Brain Res. 613 (1993) 88, Brain Res. 630 (1993) 41]. While attempting to clarify the role of the AMPA and kainate (KA) receptor subtypes in glutamatergic feeding systems, we discovered that lateral hypothalamic (LH) injection of high doses of the competitive AMPA/KA receptor antagonist, NBQX (10 and 30 nmol), elicited a pronounced feeding response. We questioned whether this effect was due to inactivation of AMPA or possibly KA receptors. To determine whether other AMPA/KA antagonists can also elicit feeding, we tested whether injection of CNQX, another AMPA/KA receptor antagonist, also stimulates eating and whether these feeding stimulatory effects were due to antagonists' actions in the LH or in other hypothalamic sites. Here we report that NBQX and CNQX elicit feeding in a dose dependent manner and are most effective when injected into the perifornical hypothalamus (PFH), or into the paraventricular nucleus (PVN) and, to a lesser extent, into the LH of satiated rats. In contrast, AMPA was most effective in stimulating feeding when injected into the LH, confirming previous reports. These data suggest that either activation or inactivation of AMPA/KA receptors in distinct but overlapping hypothalamic sites may be sufficient to induce feeding behavior, indicating a broadened role for glutamate in hypothalamic feeding mechanisms.

Regional differences in feeding and other behaviors elicited by N-methyl-D-aspartic acid in the rodent hypothalamus: a reverse microdialysis mapping study.

Regional differences in the feeding stimulatory actions of hypothalamically delivered N-methyl-D-aspartate (NMDA) were investigated. NMDA (660 microM intraprobe) delivered by reverse microdialysis into the tuberal lateral hypothalamus (tLH) reliably elicited feeding in satiated rats. The average food intake was 8.6 g in 50 min, and during the infusion rats spent 26% of the time eating, compared to less than 1% before NMDA treatment. In contrast, NMDA did not affect feeding when reverse dialyzed into the anterior LH (aLH), posterior LH (pLH) or the medial hypothalamus (MH). NMDA had no apparent behavioral effect in the aLH; in contrast, it significantly decreased the time spent resting/sleeping when infused into each of the other three areas tested. Additionally, in the medial hypothalamus, NMDA infusions increased time spent grooming; while in the pLH only alertness was significantly increased. These data underscore the functional and anatomical heterogeneity of the hypothalamus, and implicate glutamate and NMDA receptors in different portions of the hypothalamus in the control of eating, grooming and arousal.

DMSO as a vehicle for central injections: tests with feeding elicited by norepinephrine injected into the paraventricular nucleus.

Dimethyl sulfoxide (DMSO) is becoming increasingly popular as a vehicle in studies employing central injections. The aim of the present study was to determine whether the vehicle required for solubilization of substances for central injection [75% DMSO and 25% artificial CSF (aCSF)] would alter the well-characterized stimulatory response to norepinephrine (NE) injected into the paraventricular nucleus (PVN) on short-term food intake. To evaluate its suitability, we compared the effects of repeated unilateral injections of NE dissolved in two different vehicles (100% aCSF or 75% DMSO, 25% aCSF), in separate groups of animals every 48 h over a 30-day period. NE (40 nmol) stimulated food intake by approximately sevenfold compared to either vehicle alone, and the stimulatory effect was similar whether aCSF or 75% DMSO was used as a vehicle. Furthermore, the NE-induced feeding did not vary in magnitude across a series of 13 tests. These results suggest that 75% DMSO is a suitable vehicle for administering NE (and likely other water-insoluble substances)in small volumes of 0.3 microl into specific brain regions.