Restriction of dietary protein in rats increases progressive-ratio motivation for protein

Low-protein diets can impact food intake and appetite, but it is not known if motivation for food is changed. In the present study, we used an operant behavioral task – the progressive ratio test – to assess whether motivation for different foods was affected when rats were maintained on a protein-restricted diet (REST, 5% protein diet) compared to non-restricted control rats (CON, 18% protein). Rats were tested either with nutritionally-balanced pellets (18.7% protein, Experiment 1) or protein-rich pellets (35% protein, Experiment 2) as reinforcers. Protein restriction increased breakpoint for protein-rich pellets, relative to CON rats, whereas no difference in breakpoint for nutritionally-balanced pellets was observed between groups. When given free access to either nutritionally-balanced pellets or protein-rich pellets, REST and CON rats did not differ in their intake. We also tested whether a previous history of protein restriction might affect present motivation for different types of food by assessing breakpoint of previously REST animals that were subsequently put on standard maintenance chow (protein-repleted rats, REPL, Experiment 2). REPL rats did not show increased breakpoint, relative to their initial encounter with protein-rich pellets while they were protein-restricted. This study demonstrates that restriction of dietary protein induces a selective increased motivation for protein-rich food, a behavior that disappears once rats are not in need of protein

Predictive and motivational factors influencing anticipatory contrast: A comparison of contextual and gustatory predictors in food restricted and free-fed rats

In anticipation of palatable food, rats can learn to restrict consumption of a less rewarding food type resulting in an increased consumption of the preferred food when it is made available. This construct is known as anticipatory negative contrast (ANC) and can help elucidate the processes that underlie binge-like behavior as well as self-control in rodent motivation models. In the current investigation we aimed to shed light on the ability of distinct predictors of a preferred food choice to generate contrast effects and the motivational processes that underlie this behavior. Using a novel set of rewarding solutions, we directly compared contextual and gustatory ANC predictors in both food restricted and free-fed Sprague-Dawley rats. Our results indicate that, despite being food restricted, rats are selective in their eating behavior and show strong contextually-driven ANC similar to free-fed animals. These differences mirrored changes in palatability for the less preferred solution across the different sessions as measured by lick microstructure analysis. In contrast to previous research, predictive cues in both food restricted and free-fed rats were sufficient for ANC to develop although flavor-driven ANC did not relate to a corresponding change in lick patterning. These differences in the lick microstructure between context- and flavor-driven ANC indicate that the motivational processes underlying ANC generated by the two predictor types are distinct. Moreover, an increase in premature port entries to the unavailable sipper – a second measure of ANC – in all groups reveals a direct influence of response competition on ANC development

Left-right dissociation of hippocampal memory processes in mice.

Left-right asymmetries have likely evolved to make optimal use of bilaterian nervous systems; however, little is known about the synaptic and circuit mechanisms that support divergence of function between equivalent structures in each hemisphere. Here we examined whether lateralized hippocampal memory processing is present in mice, where hemispheric asymmetry at the CA3-CA1 pyramidal neuron synapse has recently been demonstrated, with different spine morphology, glutamate receptor content, and synaptic plasticity, depending on whether afferents originate in the left or right CA3. To address this question, we used optogenetics to acutely silence CA3 pyramidal neurons in either the left or right dorsal hippocampus while mice performed hippocampus-dependent memory tasks. We found that unilateral silencing of either the left or right CA3 was sufficient to impair short-term memory. However, a striking asymmetry emerged in long-term memory, wherein only left CA3 silencing impaired performance on an associative spatial long-term memory task, whereas right CA3 silencing had no effect. To explore whether synaptic properties intrinsic to the hippocampus might contribute to this left-right behavioral asymmetry, we investigated the expression of hippocampal long-term potentiation. Following the induction of long-term potentiation by high-frequency electrical stimulation, synapses between CA3 and CA1 pyramidal neurons were strengthened only when presynaptic input originated in the left CA3, confirming an asymmetry in synaptic properties. The dissociation of hippocampal long-term memory function between hemispheres suggests that memory is routed via distinct left-right pathways within the mouse hippocampus, and provides a promising approach to help elucidate the synaptic basis of long-term memory.http://dx.doi.org/10.1073/pnas.1405648111This is the author's accepted manuscript. It will be under embargo for 6 months from the date of publication. The final version is available from PNAS at http://www.pnas.org/content/early/2014/09/16/1405648111.abstract

Nicotinic α4 Receptor-Mediated Cholinergic Influences on Food Intake and Activity Patterns in Hypothalamic Circuits.

Nicotinic acetylcholine receptors (nAChRs) play an important role in regulating appetite and have been shown to do so by influencing neural activity in the hypothalamus. To shed light on the hypothalamic circuits governing acetylcholine's (ACh) regulation of appetite this study investigated the influence of hypothalamic nAChRs expressing the α4 subunit. We found that antagonizing the α4β2 nAChR locally in the lateral hypothalamus with di-hydro-ß-erythroidine (DHβE), an α4 nAChR antagonist with moderate affinity, caused an increase in food intake following free access to food after a 12 hour fast, compared to saline-infused animals. Immunocytochemical analysis revealed that orexin/hypocretin (HO), oxytocin, and tyrosine hydroxylase (TH)-containing neurons in the A13 and A12 of the hypothalamus expressed the nAChR α4 subunit in varying amounts (34%, 42%, 50%, and 51%, respectively) whereas melanin concentrating hormone (MCH) neurons did not, suggesting that DHβE-mediated increases in food intake may be due to a direct activation of specific hypothalamic circuits. Systemic DHβE (2 mg/kg) administration similarly increased food intake following a 12 hour fast. In these animals a subpopulation of orexin/hypocretin neurons showed elevated activity compared to control animals and MCH neuronal activity was overall lower as measured by expression of the immediate early gene marker for neuronal activity cFos. However, oxytocin neurons in the paraventricular hypothalamus and TH-containing neurons in the A13 and A12 did not show differential activity patterns. These results indicate that various neurochemically distinct hypothalamic populations are under the influence of α4β2 nAChRs and that cholinergic inputs to the lateral hypothalamus can affect satiety signals through activation of local α4β2 nAChR-mediated transmission.This work was supported by the Royal Society and the European Union (Latin America/European Liason, LAEL).This is the final version of the article. It first appeared from PLOS via http://dx.doi.org/1371/journal.pone.013332

Optogenetic probing of fast glutamatergic transmission from hypocretin/orexin to histamine neurons in situ

Hypothalamic hypocretin/orexin (hcrt/orx) neurons coordinate sleep-wake cycles, reward seeking, and body energy balance. Neuro-chemical data suggest that hcrt/orx cells contain several transmitters, but what hcrt/orx cells release onto their projection targets is unknown. A major pathway by which hcrt/orx neurons are thought to promote arousal is through projections to tuberomammillary histamine (HA) neurons. To study the impact of the electrical activity in hcrt/orx cells on HA neurons, we genetically targeted the light-activated excitatory ion channel channelrhodopsin-2 (ChR2) to the plasma membrane of hcrt/orx cells, and performed patch-clamp recordings from HA cells in acute mouse brain slices. Stimulation of ChR2-containing fibers with millisecond flashes of blue light produced fast postsynaptic currents in HA neurons, with ahigh connection probability (≈60% of HA cells were connected to ≈40% of hcrt/orx cells expressing ChR2). These inputs depended on tetrodotoxin-sensitive action potentials, had kinetics typical of glutamatergic responses mediated by AMPA receptors, were blocked by the AMPA receptor blocker CNQX, and displayed multiple forms of short-term plasticity (depression in ≈70% trials, facilitation in ≈30% trials, both often in the same cell). Furthermore, stimulation of hcrt/orx axons at physiological frequencies rapidly and reversibly increased action potential firing in HA cells, an effect that was abolished by blockade of AMPA receptors. These results provide the first functional evidence that hcrt/orx neurons are capable of fast glutamatergic control of their projection targets, and suggest that variations in electrical activity of hcrt/orx axons can induce rapid changes in long-range signals generated by HA neurons. ©2012 the authors

Accumbal Cholinergic Interneurons Differentially Influence Motivation Related to Satiety Signaling

Satiety, rather than all or none, can instead be viewed as a cumulative decrease in the drive to eat that develops over the course of a meal. The nucleus accumbens (NAc) is known to play a critical role in this type of value reappraisal, but the underlying circuits that influence such processes are unclear. Although NAc cholinergic interneurons (CINs) comprise only a small proportion of NAc neurons, their local impact on reward-based processes provides a candidate cell population for investigating the neural underpinnings of satiety. The present research therefore aimed to determine the role of NAc-CINs in motivation for food reinforcers in relation to satiety signaling. Through bidirectional control of CIN activity in mice, we show that when motivated by food restriction, increasing CIN activity led to a reduction in palatable food consumption while reducing CIN excitability enhanced food intake. These activity-dependent changes developed only late in the session and were unlikely to be driven by the innate reinforcer strength, suggesting that CIN modulation was instead impacting the cumulative change in motivation underlying satiety signaling. We propose that on a circuit level, an overall increase in inhibitory tone onto NAc output neurons played a role in the behavioral results, as activating NAc-CINs led to an inhibition of medium spiny neurons that was dependent on nicotinic receptor activation. Our results reveal an important role for NAc-CINs in controlling motivation for food intake and additionally provide a circuit-level framework for investigating the endogenous cholinergic circuits that signal satiety.Peer reviewe

Optogenetic evidence for inhibitory signaling from orexin to MCH neurons via local microcircuits.

The lateral hypothalamus (LH) is a key regulator of multiple vital behaviors. The firing of brain-wide-projecting LH neurons releases neuropeptides promoting wakefulness (orexin/hypocretin; OH), or sleep (melanin-concentrating hormone; MCH). OH neurons, which coexpress glutamate and dynorphin, have been proposed to excite their neighbors, including MCH neurons, suggesting that LH may sometimes coengage its antagonistic outputs. However, it remains unclear if, when, and how OH actions promote temporal separation of the sleep and wake signals, a process that fails in narcolepsy caused by OH loss. To explore this directly, we paired optogenetic stimulation of OH cells (at rates that promoted awakening in vivo) with electrical monitoring of MCH cells in mouse brain slices. Membrane potential recordings showed that OH cell firing inhibited action potential firing in most MCH neurons, an effect that required GABAA but not dynorphin receptors. Membrane current analysis showed that OH cell firing increased the frequency of fast GABAergic currents in MCH cells, an effect blocked by antagonists of OH but not dynorphin or glutamate receptors, and mimicked by bath-applied OH peptide. In turn, neural network imaging with a calcium indicator genetically targeted to MCH neurons showed that excitation by bath-applied OH peptides occurs in a minority of MCH cells. Collectively, our data provide functional microcircuit evidence that intra-LH feedforward loops may facilitate appropriate switching between sleep and wake signals, potentially preventing sleep disorders.This work was supported by the Royal Society Dorothy Hodgkin Fellowship (JAS) and HFSP 37 Young Investigator Award (DB and AA, award ref. RGY0076/2012).This is the final version. It first appeared at http://www.jneurosci.org/content/35/14/5435.abstract

Sex difference in physical activity, energy expenditure and obesity driven by a subpopulation of hypothalamic POMC neurons.

OBJECTIVE: Obesity is one of the primary healthcare challenges of the 21st century. Signals relaying information regarding energy needs are integrated within the brain to influence body weight. Central among these integration nodes are the brain pro-opiomelanocortin (POMC) peptides, perturbations of which disrupt energy balance and promote severe obesity. However, POMC neurons are neurochemically diverse and the crucial source of POMC peptides that regulate energy homeostasis and body weight remains to be fully clarified. METHODS: Given that a 5-hydroxytryptamine 2c receptor (5-HT2CR) agonist is a current obesity medication and 5-HT2CR agonist's effects on appetite are primarily mediated via POMC neurons, we hypothesized that a critical source of POMC regulating food intake and body weight is specifically synthesized in cells containing 5-HT2CRs. To exclusively manipulate Pomc synthesis only within 5-HT2CR containing cells, we generated a novel 5-HT 2C R (CRE) mouse line and intercrossed it with Cre recombinase-dependent and hypothalamic specific reactivatable Pomc (NEO) mice to restrict Pomc synthesis to the subset of hypothalamic cells containing 5-HT2CRs. This provided a means to clarify the specific contribution of a defined subgroup of POMC peptides in energy balance and body weight. RESULTS: Here we transform genetically programed obese and hyperinsulinemic male mice lacking hypothalamic Pomc with increased appetite, reduced physical activity and compromised brown adipose tissue (BAT) into lean, healthy mice via targeted restoration of Pomc function only within 5-HT2CR expressing cells. Remarkably, the same metabolic transformation does not occur in females, who despite corrected feeding behavior and normalized insulin levels remain physically inactive, have lower energy expenditure, compromised BAT and develop obesity. CONCLUSIONS: These data provide support for the functional heterogeneity of hypothalamic POMC neurons, revealing that Pomc expression within 5-HT2CR expressing neurons is sufficient to regulate energy intake and insulin sensitivity in male and female mice. However, an unexpected sex difference in the function of this subset of POMC neurons was identified with regard to energy expenditure. We reveal that a large sex difference in physical activity, energy expenditure and the development of obesity is driven by this subpopulation, which constitutes approximately 40% of all POMC neurons in the hypothalamic arcuate nucleus. This may have broad implications for strategies utilized to combat obesity, which at present largely ignore the sex of the obese individual