Leptin facilitates learning and memory performance and enhances hippocampal CA1 long-term potentiation and CaMK II phosphorylation in rats.

Leptin, an adipocytokine encoded by an obesity gene and expressed in adipose tissue, affects feeding behavior, thermogenesis, and neuroendocrine status via leptin receptors distributed in the brain, especially in the hypothalamus. Leptin may also modulate the synaptic plasticity and behavioral performance related to learning and memory since: leptin receptors are found in the hippocampus, and both leptin and its receptor share structural and functional similarities with the interleukin-6 family of cytokines that modulate long-term potentiation (LTP) in the hippocampus. We therefore examined the effect of leptin on (1) behavioral performance in emotional and spatial learning tasks, (2) LTP at Schaffer collateral-CA1 synapses, (3) presynaptic and postsynaptic activities in hippocampal CA1 neurons, (4) the intracellular Ca(2+) concentration ([Ca(2+)](i)) in CA1 neurons, and (5) the activity of Ca(2+)/calmodulin protein kinase II (CaMK II) in the hippocampal CA1 tissue that exhibits LTP. Intravenous injection of 5 and/or 50mug/kg, but not of 500mug/kg leptin, facilitated behavioral performance in passive avoidance and Morris water-maze tasks. Bath application of 10(-12)M leptin in slice experiments enhanced LTP and increased the presynaptic transmitter release, whereas 10(-10)M leptin suppressed LTP and reduced the postsynaptic receptor sensitivity to N-methyl-d-aspartic acid. The increase in the [Ca(2+)](i) induced by 10(-10)M leptin was two times greater than that induced by 10(-12)M leptin. In addition, the facilitation (10(-12)M) and suppression (10(-10)M) of LTP by leptin was closely associated with an increase and decrease in Ca(2+)-independent activity of CaMK II. Our results show that leptin not only affects hypothalamic functions (such as feeding, thermogenesis, and neuroendocrine status), but also modulates higher nervous functions, such as the behavioral performance related to learning and memory and hippocampal synaptic plasticity.

Impairment of long-term potentiation and spatial memory in leptin receptor-deficient rodents.

Leptin is well known to be involved in the control of feeding, reproduction and neuroendocrine functions through its action on the hypothalamus. However, leptin receptors are found in brain regions other than the hypothalamus (including the hippocampus and cerebral cortex) suggesting extrahypothalamic functions. We investigated hippocampal long-term potentiation (LTP) and long-term depression (LTD), and the spatial-memory function in two leptin receptor-deficient rodents (Zucker rats and db/db mice). In brain slices, the CA1 hippocampal region of both strains showed impairments of LTP and LTD; leptin (10(-12) M) did not improve these impairments in either strain. These strains also showed lower basal levels of Ca(2+)/calmodulin-dependent protein kinase II activity in the CA1 region than the respective controls, and the levels did not respond to tetanic stimulation. These strains also showed impaired spatial memory in the Morris water-maze test (i.e. longer swim-path lengths during training sessions and less frequent crossings of the platform's original location in the probe test. From these results we suggest that the leptin receptor-deficient animals show impaired LTP in CA1 and poor spatial memory due, at least in part, to a deficiency in leptin receptors in the hippocampus.

Orexin-A (hypocretin-1) impairs Morris water maze performance and CA1-Schaffer collateral long-term potentiation in rats.

Glucose-sensitive neurons in the lateral hypothalamic area produce orexin-A (hypocretin-1) and orexin-B (hypocretin-2) and send their axons to the hippocampus, which predominantly expresses orexin receptor 1 showing a higher sensitivity to orexin-A. The purpose of the present study was to assess the effects of orexin-A on the performance of Wistar rats during the Morris water maze test and then to determine the effects of orexin-A on both the long-term potentiation and long-term depression in Schaffer collateral/commissural-CA1 synapses in hippocampal slices. The results of the Morris water maze test show that 1.0 and 10 nmol of orexin-A, when administered intracerebroventricularly, retarded spatial learning. A probe test examined after training of water maze task also showed an impairment in spatial memory. The results of an electrophysiological study using hippocampal slices demonstrated that 1.0 to 30 nM of orexin-A applied to the perfusate produces a dose-dependent and time dependent suppression of the long-term potentiation. In addition, the long-term depression was not affected by orexin-A. The results of a paired-pulse facilitation experiment indicated that the effects of orexin-A were post-synaptic and not due to presynaptic transmitter release. These results show that orexin-A impairs spatial performance and these impairments can be attributed to a suppression of long-term potentiation in the Schaffer collateral-CA1 hippocampal synapses.

Pain modulatory actions of cytokines and prostaglandin E2 in the brain.

Proinflammatory cytokines such as IL-1, IL-6, and TNF alpha are known to enhance nociception at peripheral inflammatory tissues. These cytokines are also produced in the brain. We found that an intracerebroventricular injection of IL-1 beta only at nonpyrogenic doses in rats reduced the paw-withdrawal latency on a hot plate and enhanced the responses of the wide dynamic range neurons in the trigeminal nucleus caudalis to noxious stimuli. This hyperalgesia, as assessed by behavioral and neuronal responses, was blocked by pretreatment with IL-1 receptor antagonist (IL-1Ra), Na salicylate, or alpha melanocyte-stimulating hormone, indicating the involvement of IL-1 receptors and the synthesis of prostanoids. IL-6 and TNF alpha at nonpyrogenic doses also induced hyperalgesia in a prostanoid-dependent way. Furthermore, the preoptic area (POA) was most sensitive to IL-1 beta (5-50 pg/kg) in the induction of behavioral hyperalgesia. The maximal response was obtained 30 min after injection of IL-1 beta at 20 pg/kg. On the other hand, an injection of IL-1 beta (20-50 pg/kg) into the ventromedial hypothalamus (VMH) prolonged the paw-withdrawal latency maximally 10 min after injection. This analgesia, as well as the intraPOA IL-1 beta-induced hyperalgesia, was completely blocked by IL-1Ra or Na salicylate. Our previous study has revealed that i.c.v. injection of PGE2 induces hyperalgesia through EP3 receptors and analgesia through EP1 receptors by its central action. The results, taken together, suggest (1) that IL-1 beta at lower doses in the brain induces hyperalgesia through EP3 receptors in the POA and (2) that the higher doses of brain IL-1 beta produces analgesia through EP1 receptors, probably, in the VMH.

Complex functional attributes of amygdaloid gustatory neurons in the rhesus monkey.

To reveal specific functions of glucose-sensitive (GS) and glucose-insensitive (GIS) cells in chemical information processing, single neuron activity was recorded in the amygdaloid body (AMY) of macaques during: 1) gustatory stimulations and 2) micro-electrophoretic administration of chemicals. Of the 629 neurons tested, 56 (8.9%) responded to, usually two or more, taste qualities. Hedonically distinct tastants usually elicited opposite firing rate changes of the gustatory cells. Seventy percent of the gustatory responses were recorded from GS neurons (17% of all AMY cells). Catecholamines (CAs) induced discharge rate changes in a majority of taste-responsive neurons: The GS gustatory cells were suppressed by norepinephrine (in the form of noradrenaline HCl, NA), whereas the GIS taste-responsive neurons were facilitated by dopamine (DA). Furthermore, NA- and/or DA-antagonists were able to attenuate or suppress taste-elicited responses of several of these cells. These and previous data indicate a specific functional organization of AMY gustatory cells: The GS and GIS taste neurons appear to be involved in differential integration of feeding-associated humoral-metabolic, motivational and exogenous chemical information.

Two-generation reproductive toxicity study of tributyltin chloride in male rats.

A 2-generation reproductive toxicity study of tributyltin chloride (TBTCl) was conducted in male rats using dietary concentrations of 5, 25, and 125 ppm TBTCl to evaluate its effect on sexual development and the reproductive system. F1 males were killed on postnatal day 119 and F2 males were killed on postnatal day 91. TBTCl affected the male reproductive system of rats. The weights of the testis and epididymis were decreased and homogenization-resistant spermatid and sperm count were reduced mainly in the 125 ppm TBTCl group. Histopathologic changes were also observed in the testis of this group and included vacuolization of the seminiferous epithelium, spermatid retention, and delayed spermiation. However, the changes were minimal in nature. The weight of the ventral prostate was decreased to 84% of the control value in the 125 ppm group in the F1 generation and decreased to 84 and 69% of the control value in the 25 ppm and 125 ppm TBTCl groups, respectively, in the F2 generation. The serum 17beta-estradiol concentration was also decreased to 55% of the control value in the 125 ppm group in the F1 generation and decreased to 78 and 57% of the control value in the 25 ppm and 125 ppm TBTCl groups, respectively, in the F2 generation. However, the serum concentrations of luteinizing hormone (LH) and testosterone were not decreased in these groups. These changes corresponded with those caused by aromatase inhibition and therefore TBTCl might be a weak aromatase inhibitor in male rats.

A computerized control system for a bar-press feeding task initiated by monkey vocalization.

Computerized control of a bar-press feeding task initiated by a monkey's vocalization was designed to study the motivational roles of brain regions in feeding behavior. In this paradigm, the monkey was required to vocalize in order to initiate the task and then press the bar 30 times to obtain the reward. A microcomputer was used to sense the vocalization and to control the task. Using this system, changes in single neuron activity in the orbitofrontal cortex and the lateral hypothalamic area preceding the time of the vocalization were observed. The entire system including interface and control program is described.

Intracerebroventricular injection of interleukin-1 beta enhances nociceptive neuronal responses of the trigeminal nucleus caudalis in rats.

To assess the effect of interleukin-1 (IL-1) in the brain on nociception electrophysiologically, recombinant human IL-1 beta (rhIL-1 beta) (1 pg/kg to 1 microgram/kg, i.e., 0.29 pg-0.33 microgram/rat) was microinjected into the lateral cerebral ventricle of urethane-anesthetized rats and the changes of responses in the wide dynamic range (WDR) neurons in the trigeminal nucleus caudalis to noxious pinching of facial skin were observed. A significant enhancement in the responses of the WDR neurons to noxious stimuli was observed after the injection of rhIL-1 beta between 10 pg/kg and 1 ng/kg, which showed a maximal response at a dose of 100 pg/kg (29-33 pg/rat) which began to appear 5 min after injection, reached a peak within 25 min and then gradually subsided. However, this dose of rhIL-1 beta did not affect the responses of low threshold mechanoreceptive neurons to skin brushing. An increase in the dose of rhIL-1 beta by more than 10 ng/kg (up to 1 microgram/kg) had no effect on the nociceptive responses of the WDR neurons. The rhIL-1 beta-induced enhancement of nociceptive responses of WDR neurons was completely abolished by pretreatment with either IL-1 receptor antagonist, Na salicylate or alpha-melanocyte stimulating hormone. These results therefore provide electrophysiological evidence that IL-1 beta which is produced in the brain induces hyperalgesia in the rat.

Intracerebroventricular injection of interleukin-1 beta induces hyperalgesia in rats.

To determine whether interleukin-1 beta (IL-1 beta) in the brain may modulate nociception, recombinant human IL-1 beta (rhIL-1 beta) (1 pg/kg to 1 microgram/kg) was microinjected into the lateral cerebral ventricle of rats and the latency before initiating the licking of their hindpaws after being placed on a hot plate (50.0 +/- 0.1 degrees C) was measured. A significant reduction of the paw-lick latency was observed after injections of nonpyrogenic doses (10 pg/kg to 1 ng/kg) of rhIL-1 beta, showing a maximal response at a dose of 100 pg/kg which began to appear 5 min after injection, reached a peak within 30 min and then gradually subsided. An increase in the amount of rhIL-1 beta to > 1 ng/kg (up to 1 microgram/kg) had no effect on the nociceptive threshold. The rhIL-1 beta-induced hyperalgesia was completely abolished by pretreatment with an IL-1 receptor antagonist (IL-1ra) or Na salicylate. Similar pretreatment with alpha-melanocyte-stimulating hormone (alpha-MSH) also inhibited the rhIL-1 beta-induced hyperalgesia. However, pretreatment with alpha-helical corticotropin-releasing factor (CRF)9-41 failed to affect it. The results suggest that IL-1 beta in the brain produces hyperalgesia by its receptor-mediated and prostaglandin-dependent action which is sensitive to alpha-MSH. The hyperalgesic action of central IL-1 does not appear to depend on the CRF system.

Neuron activity of the ventromedial hypothalamus and the medial preoptic area of the female monkey during sexual behavior.

To elucidate the neural mechanisms of the sexual behavior of the female monkey, single neuron activity of the ventromedial hypothalamus (VMH) and the medial preoptic area (MPOA) was recorded during sexual behavior with a male partner. Copulation was initiated when either the male's touch elicited passive presenting ('receptive presenting') or when the female's active presenting ('proceptive presenting') evoked mounting from the male. Proceptive presenting with no mating acts evoked activity changes in about 40% of cells in the VMH (mainly excitation) and MPOA (mainly inhibition). When the male's mating acts were manifested, 56% of VMH cells and 87% of MPOA cells showed firing changes during presenting, with the proportion of MPOA cells which showed excitation significantly increased. Progress of mating acts of the male partner (e.g. increases in the rate and/or number of thrusting), enhanced activity changes in about 40% of VMH and 70% of MPOA cells tested. The findings suggest that VMH and MPOA cells are involved in the control of sexual behavior in different ways: excitation of the VMH and inhibition of MPOA are related to execution of presenting, while excitation of MPOA is related to sexual intercourse with a male partner; and mating acts of a male partner modulate activity of both VMH and MPOA neurons of the female.

Medial preoptic and hypothalamic neuronal activity during sexual behavior of the male monkey.

Neuronal activity changes in the medial preoptic area of the male monkey were related to the commencement of sexual behavior, penile erection and the refractory period following ejaculation. Increased neuronal activity in the dorsomedial hypothalamic nucleus was found to be synchronized to each mating act. The involvement of medial preoptic neurons in sexual arousal, initial penile erection and that of dorsomedial hypothalamic neurons in the copulatory act are suggested by the present findings.

Influence of acetylcholine on neuronal activity in monkey orbitofrontal cortex during bar press feeding task.

Cholinergic involvement in feeding-related neuronal activity was investigated in the orbitofrontal cortex (OBF) of the behaving monkey by means of a microiontophoretic application of acetylcholine (ACh) and atropine. The activity of ACh sensitive cells did not correlate with any particular event during the bar press feeding task. Atropine blocked the excitatory response to ACh and task events. The results suggest that the cholinergic system in the monkey OBF regulates excitatory neuronal responses in several phases of motivated behavior.

Intracerebroventricular injection of prostaglandin E2 induces thermal hyperalgesia in rats: the possible involvement of EP3 receptors.

To determine what types of prostanoid receptors are involved in the central effect of prostaglandin E2 (PGE2) on nociception, we administered PGE2 and its agonists, i.e., 17-phenyl-omega-trinor PGE2 (an EP1 receptor agonist), butaprost (an EP2 receptor agonist), 11-deoxy PGE1 (an EP2/EP3 receptor agonist, EP2 >> EP3) and M&B28767 (an EP3 receptor agonist) into the lateral cerebroventricle (LCV) of rats and observed the changes of paw-withdrawal latency on a hot plate. The LCV injection of PGE2 (10 pg/kg-10 ng/kg), 11-deoxy PGE1 (100 pg/kg-10 ng/kg) and M&B28767 (1 pg/kg-100 pg/kg) produced a significant reduction in the paw-withdrawal latency. The maximal reduction was observed 15 min after the LCV injection of these drugs. Neither 17-phenyl-omega-trinor PGE2 (1 pg/kg-1 microgram/kg) nor butaprost (1 pg/kg-100 microgram/kg) induced any significant changes in the paw-withdrawal latency. The LCV injection of PGE2 (1 microgram/kg) and 17-phenyl-omega-trinor PGE2 (50 micrograms/kg) increased the latency only 5 min after LCV injection. These findings indicate that the LCV injection of PGE2 induces thermal hyperalgesia through EP3 receptors and analgesia through EP1 receptors by its central action in rats.

Identification of differently timed motor components of conditioned blink responses.

Electromyographic recordings were made from the orbicularis oculi muscles of cats in order to identify differently timed motor components of conditioned eye blink responses (CRs). Conditioning was established rapidly by pairing electrical stimulation of the hypothalamus (HS) with a click conditioned stimulus (CS) and a glabella tap unconditioned stimulus (US). Analysis of the EMG responses disclosed five different motor components of the CR that could be distinguished and characterized according to their latencies of occurrence. Four were associated with an increase in EMG activity elicited by the CS (16-48 ms, alpha(1); 48-80 ms, alpha(2); 80 to 120 ms, beta; >/=120 ms, gamma), and one was associated with a decrease in activity (16 to 60 ms, alpha(i)). Analysis of the amplitudes of the different components of the CR during the course of conditioning and extinction disclosed that short latency, alpha(1) components of the CRs were acquired and extinguished in a manner equivalent to longer latency components of the CRs. The observations supported the hypothesis that short and long latency components of blink responses represented comparable rather than substantially different forms of Pavlovian conditioning. The alpha(2) response was present before conditioning began, and increased with other components after conditioning. The alpha(i) response component was also observed prior to conditioning, and represents a previously undetected, inhibitory consequence of presenting weak (70 dB) acoustic stimuli. It could play a role in conditioned inhibition, latent inhibition and blocking as well as suppression of the conditioned motor response during extinction.

Catecholaminergic mechanisms of feeding-related lateral hypothalamic activity in the monkey.

The functional role of the catecholaminergic mechanism in the lateral hypothalamus (LHA), in feeding behavior of the monkey was investigated by single neuron activity recording and electrophoretic application of dopamine (DA), noradrenaline (NA) and their antagonists. The feeding paradigm had 4 phases: cue light (CL) signaled start of bar press; bar press (BP, 20-30 times); short cue tone (CT) triggered by last bar press signaled presentation of food; and ingestion-reward (RW). Of 312 neurons tested, 189 (61%) responded in one or more phases of the feeding task. Two types of response were observed: CL- or CT-related transient, and BP- or RW-related long-lasting responses. These feeding-related responses depended on the nature of the food and on the hunger-satiety level. DA excited or inhibited different neurons, while NA mainly inhibited firing. DA-sensitive neurons responded more often in the feeding task than insensitive neurons due mainly to differences in responsiveness to CL on (chi 2 test, P less than 0.01), at motor initiation, and during BP (P less than 0.05). Spiperone blocked the former two responses. NA-sensitive neurons responded more often in the feeding task due to responsiveness during BP and RW (P less than 0.01). Sotalol blocked some BP-related responses, and phenoxybenzamine and sotalol blocked the CT-related responses. The data suggest that dopaminergic and noradrenergic inputs in the LHA are crucial in task initiation and reward processing, respectively. Integration of these catecholaminergic and other inputs in the LHA might be important in accomplishing motivated feeding.

Proceptive presenting elicited by electrical stimulation of the female monkey hypothalamus.

Proceptive presenting by female macaque monkeys was evoked by electrical stimulation of the ventromedial hypothalamic nucleus and the medial preoptic area, under conditions of partial restraint while sitting in a primate chair. This behavior could be elicited only when a male monkey was in close proximity and not when he was removed or was replaced with a female monkey or the human experimenter. This seems to be the first report on the effects of electrical brain stimulation on proceptivity in the female monkey.

Opiate mechanism in reward-related neuronal responses during operant feeding behavior of the monkey.

Reward-related neuronal activity and its modulation by morphine and naloxone was investigated by extracellular single neuron recording and electrophoretic application of drugs in the lateral hypothalamus, during operant feeding of the monkey. Morphine-sensitive neurons responded more often during bar press and ingestion-reward phases. Naloxone blocked only ingestion-reward responses, especially the inhibitory ones. The results suggest that the central opiate system can be involved in reward-related neuronal responses in the lateral hypothalamic area of the monkey.

Effects of behaviorally rewarding hypothalamic electrical stimulation on intracellularly recorded neuronal activity in the motor cortex of awake monkeys.

Effects of hypothalamic stimulation (HS) were studied in intracellular recordings obtained from 125 neurons of the motor cortex (MC). HS that was effective in reinforcing bar-press behavior, i.e. satisfactory for intracranial self-stimulation (ICSS), evoked short-latency (less than 3 ms) activation of these cortical neurons more frequently (42% of cells tested) than did HS that was ineffective in reinforcing bar-press behavior (7% of cells tested). Longer latency activation (greater than 3 ms) and inhibition (of variable onset) also occurred, but their incidence was not significantly different when HS was effective or ineffective in producing ICSS. Effects of HS that was effective in producing ICSS were also examined in 23 cells in which the spikes were followed by afterhyperpolarization (AHP) of 1.4-10 mV amplitude and 1.7-54 ms duration. The amplitudes of AHPs of greater than 8 ms duration were reduced after presentations of HSs that were effective as a reinforcer for ICSS. These results suggest that: (1) MC neurons receive reward-related hypothalamic information through pathways sufficiently direct to produce short-latency activation; and (2) a modulation of spike afterhyperpolarization can be observed in conjunction with reception of this information.

Influence of acetylcholine on neuronal activity of monkey amygdala during bar press feeding behavior.

Single neuron activity in the monkey amygdala was investigated during cue signalled conditioned bar press feeding behavior and the effects of electrophoretically applied acetylcholine (ACh) and atropine were analyzed. ACh increased the firing rate of one third of the neurons tested; these excitatory responses were inhibited by the muscarinic receptor antagonist atropine. No characteristic location of ACh-sensitive neurons was found, cells were diffusely distributed throughout the amygdala. Activity of ACh-sensitive neurons did not correlate with any particular event during the bar press feeding task. However, continuous application of ACh at low current intensity during the task significantly enhanced the task-related excitatory firing patterns, or markedly attenuated the inhibitory responses. Continuous application of atropine elicited or enhanced inhibitory response patterns. These results suggest that the cholinergic system of the monkey amygdala facilitates neuronal excitation but attenuates inhibition related to various phases of feeding behavior, such as to cue recognition, food aquisition and rewarding process.

The opposing effects of interleukin -1 beta microinjected into the preoptic hypothalamus and the ventromedial hypothalamus on nociceptive behavior in rats.

The effects of microinjections of recombinant human interleukin-1 beta (rhIL-1 beta) into the hypothalamus and neighboring basal forebrain on nociceptive behavior were studied using a hot-plate test in rats. The microinjection of rhIL-1 beta at doses between 5 pg/kg and 50 pg/kg into the medial part of the preoptic area (MPO) reduced the paw-withdrawal latency. The maximal reduction was obtained 30 min after the injection of rhIL-1 beta at 20 pg/kg. RhIL-1 beta (20 pg/kg)-induced hyperalgesia was completely blocked by the simultaneous injection of IL-1 receptor antagonist (IL-1ra, 20 ng/kg), Na salicylate (200 ng/kg) or alpha-melanocyte-stimulating hormone alpha-MSH, 20 ng/kg). The intra-MPO injection of rhIL-1 beta at doses of less than 5 pg/kg or more than 50 pg/kg (up to 2 ng/kg) into the paraventricular nucleus, the lateral hypothalamic area and the septal nucleus had no effect on nociception. The microinjection rhIL-1 beta (20 pg/kg-50 pg/kg) into the ventromedial hypothalamus produced a prolongation of the paw-withdrawal latency. A maximal prolongation was obtained 10 min after the injection of rhIL-1 beta at 50 pg/kg. This reaction was also blocked by the simultaneous injection of IL-1ra (50 ng/kg) and Na salicylate (500 ng/kg). These findings indicate that IL-1 beta in the MPO and the VMH produces hyperalgesia and analgesia, respectively, while, in addition, both effects are mediated by IL-1 receptors and the synthesis of prostaglandins.