Matthew J. Wayner

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.

Orexin-A (Hypocretin-1) and leptin enhance LTP in the dentate gyrus of rats in vivo

Orexin-A (Hypocretin-1) has been localized in the posterior and lateral hypothalamic perifornical region. Orexin containing axon terminals have been found in hypothalamic nuclei and many other parts of the brain; for example, the hippocampus. Two types of orexin receptors have been discovered. Orexin 1 type of receptors have been described and been shown to be widely distributed in the rat brain including the hippocampus. Subsequently Orexin-A was found to impair both water maze performance and hippocampal long term potentiation (LTP). Leptin is expressed in adipose tissue and released into the blood where it affects food intake and can also produce widespread physiological changes mediated via autonomic preganglionic neurons, pituitary gland, and cerebral cortex. Immunoreactivity for leptin receptors has been found in various hypothalamic nuclei including the lateral hypothalamic area as well as the hippocampus especially in the dentate gyrus and CA1. Leptin receptor deficient rats and mice also show impaired LTP in CA1 and poor performance in the water maze. The present study was conducted to determine the effects of 0.0, 30, 60, 90, and 100 nM, orexin-A, and leptin, 0.0, 1.0, 100 nM, 1, and 10 microM, in 1.0 microl of ACSF, applied directly into the dentate gyrus, on LTP in medial perforant path dentate granule cell synapses in urethane anesthetized rats. Orexin-A specifically enhanced LTP at the 90 nM dose; and it was possible to block the enhancement by pretreating the animals with SB-334867, a specific orexin 1 receptor antagonist. Leptin enhanced normal LTP at 1.0 microM but inhibited LTP at lower and higher doses. These results and previous data indicate that the same peptide could possibly have different modulatory post synaptic effects in different hippocampal synapses dependent upon different types of post synaptic receptors.

Effects of leptin and orexin-A on food intake and feeding related hypothalamic neurons

The lateral hypothalamic area (LHA) and the ventromedial hypothalamic nucleus (VMH) have historically been implicated in ingestive behavior, energy balance and body mass regulation. The LHA is more closely associated with the initiation of eating; whereas the VMH mediates the cessation of eating. The parvocellular part of the paraventricular nucleus (pPVN) is also included in the suppressing mechanism. Recently, two hypothalamic peptides, orexin-A and orexin-B, localized in the posterior and lateral hypothalamic perifornical region were discovered in the rat brain and they increase food intake. Leptin, a protein encoded by an obesity gene, expressed in adipose tissue and released into the blood also affects food intake. Orexin and leptin receptors have been localized in the LHA, pPVN, and VMH. The purpose of this study was to measure food intake in the rat in response to leptin and orexin-A; and to determine their electrophysiological effects on feeding related hypothalamic neurons. Results clearly show that leptin suppresses food intake whereas orexin-A increases food intake. These differences are associated with leptin and orexin-A modulatory effects on LHA, pPVN, and VMH glucose responding neurons. In the LHA, leptin inhibits a larger proportion of both glucose-sensitive neurons (GSNs) and non-GSNs. In the pPVN, leptin increases more GSNs in comparison to non-GSNs. Whereas in the VMH, leptin increases the activity of glucoreceptor neurons (GRNs) in comparison to non-GRNs. Orexin-A had opposite effects: increases activity of GSNs more than the non-GSNs in the LHA and significantly suppresses GRNs in the VMH. In the pPVN, orexin-A had no observable effects on neurons that have a low density of orexin 2 receptors. Results are discussed in terms of hypothalamic neural circuits that are sensitive to endogenous food intake inducing and reducing substances.

Calpain inhibitors block long-term potentiation

Long-term potentiation (LTP) is a form of synaptic plasticity that serves as a model for certain types of learning and memory. The role of the calcium-activated thiol proteases or calpains in the biochemical mechanism of LTP has been explored. We show that the extracellular application of two newly developed, highly potent calpain inhibitors, N-acetyl-Leu-Leu-norleucinal and N-acetyl-Leu-Leu-methioninal, block LTP in both the Schaffer collateral-CA1 synaptic zone of the rat hippocampal slice and in perforant path-stimulated dentate granule cells in the intact hippocampus. The inhibitors do not affect baseline synaptic transmission and block LTP in the slice preparation if applied before but not after tetanic stimulation. The calpain inhibitor leupeptin is less potent than the above peptides but also blocks LTP if applied at a sufficient concentration.

Corticotrophin-releasing factor produces a long-lasting enhancement of synaptic efficacy in the hippocampus

We have previously demonstrated that intra‐hippocampal injection of corticotrophin‐releasing factor improved memory retention of an inhibitory avoidance learning in rats; while the electrophysiological effects corticotrophin‐releasing factor produces on hippocampal neurons are largely uncharacterized. In the present study, we found that corticotrophin‐releasing factor injected into the dentate gyrus of hippocampus produced a dose‐dependent and long‐lasting enhancement in synaptic efficacy of these neurons, as measured by an increase in the amplitude and slope of population excitatory postsynaptic potentials, as well as the amplitude of population spike. The onset of corticotrophin‐releasing factor‐induced potentiation was slow. It was observed approximately 40–60 min after corticotrophin‐releasing factor administration and lasted for more than 5 h. This effect of corticotrophin‐releasing factor was blocked by pretreatment with the cyclase‐adenosine‐3,5‐monophosphate (cAMP) inhibitor Rp‐adenosine‐3,5‐cyclic monophosphothiolate triethylamine (Rp‐cAMPS) and partially blocked by the N‐methyl‐D‐aspartate receptor antagonist MK‐801. Further, pretreatment with corticotrophin‐releasing factor receptor antagonist dose‐dependently diminished tetanization‐induced long‐term potentiation, and corticotrophin‐releasing factor and tetanic stimuli had an additive effect on hippocampal neuron excitation. Moreover, direct injection of corticotrophin‐releasing factor increased cAMP level in the dentate gyrus. These results together suggest that corticotrophin‐releasing factor‐induced potentiation simulates the late phase of tetanization‐induced long‐term potentiation and cAMP seems to be the messenger mediating this effect. Moreover, corticotrophin‐releasing factor‐induced potentiation and long‐term potentiation may share some similar mechanisms, and corticotrophin‐releasing factor is probably involved in the neural circuits underlying long‐term potentiation. Thus, corticotrophin‐releasing factor may play an important role in modulating synaptic plasticity in the hippocampus.

The effects of angiotensin IV analogs on long-term potentiation within the CA1 region of the hippocampus in vitro.

Within the brain-renin angiotensin system, it is generally assumed that angiotensin peptide fragments shorter than angiotensins II and III, including angiotensin IV (AngIV), are inactive. This belief has been challenged by the recent discovery that AngIV, and AngIV-like analogs, bind with high affinity and specificity to a putative angiotensin binding site termed AT4. In the brain these sites include the hippocampus, cerebellum, and cerebral cortex, and influence associative and spatial learning tasks. The present study investigated the effects of two AngIV analogs, Nle1-AngIV (an AT4 receptor agonist) and Nle1-Leual3-AngIV (an AT4 receptor antagonist), on long-term potentiation (LTP). Field excitatory postsynaptic potentials (fEPSPs) were recorded from the CA1 stratum radiatum following stimulation of the Schaffer collateral pathway. Activation of AT4 receptors by Nle1-AngIV enhanced synaptic transmission during low-frequency test pulses (0.1 Hz), and increased the level of tetanus-induced LTP by 63% over that measured under control conditions. Paired stimulation before and during infusion of Nle1-AngIV indicated no change in paired-pulse facilitation (PPF) as a result of AT4 receptor activation suggesting that the underlying mechanism(s) responsible for Nle1-AngIV-induced increase in synaptic transmission and LTP is likely a postsynaptic event. Further, applications of Nle1-Leual3-AngIV prior to, but not 15 or 30 min after, tetanization prevented stabilization of LTP. These results extend previous findings from behavioral data in that AT4 receptor agonists and antagonists are capable of activating, and inhibiting, learning and memory pathways in the hippocampus, and suggest that the AT4 receptor subtype is involved in synaptic plasticity.

Vagus nerve afferent and efferent innervation of the rat uterus : an electrophysiological and HRP study

To determine a possible brainstem connection with the uterus, a study with electrophysiological techniques and horseradish peroxidase (HRP) tracing was performed in the rat. Neurons of the nucleus of the tractus solitarius decreased in discharge frequency during cervicovaginal distension. HRP injections into the uterine walls resulted in the appearance of labelled cells in the nodose ganglion and in the dorsal motor nucleus of the vagus nerve. The results demonstrate a direct bidirectional vagal complex-uterus connection via the vagus nerve. Results are discussed in terms of a complex uterus control system in which the paraventricular nucleus might play an integrative role.

Angiotensin IV enhances LTP in rat dentate gyrus in vivo

Angiotensins have been shown to play a significant role in a variety of physiological functions including learning and memory processes. Relatively recent evidence supports the increasing importance of angiotensin IV (Ang IV), in many of these functions previously associated only with Ang II, including learning and memory. An interesting hypothesis generated by these results has been that Ang II is a precursor for the production of a more active peptide fragment, Ang IV. Since Ang II impairs learning and memory, when administered directly or released into the hippocampal dentate gyrus, and inhibits long term potentiation (LTP) in medial perforant path-dentate granule cell synapses, as well; it remained to be seen what effects Ang IV had on LTP in these same synapses. Results of this study show clearly that Ang IV significantly enhances LTP, and the enhancement is both dose and time dependent. The following solutions of Ang IV were administered over a five min period, at the end of baseline and before the first tetanus was applied: 2.39, 4.78, and 9.56 nM. An inverted U-type dose related effect was observed. A complex time related effect was observed with a maximum at 5 min, a return to normal LTP at 30 min and a minimum below normal at 90 min, and a return to normal LTP at 120 min. The effects of the 4.78 nM solution were determined at the following intervals between administration and the first tetanus: 5, 15, 30, 60, 90, and 120 min. The enhancement of LTP can be prevented by pretreatment with Divalinal, an Ang IV antagonist, without any effect on normal LTP. Two solutions of Divalinal were used; 5 nM and 5 microM, and the 5 microM was more effective and completely blocked the enhancement of normal LTP. Results were also obtained with 4.78 nM Nle1-Ang IV (Norleucine), an Ang IV agonist. Norleucine was less effective than Ang IV in the enhancement of normal LTP and displayed a similar time course of activity. Both Ang IV and Norleucine produced a significant suppression of normal LTP at 90 min; that remains to be explained. However, the inhibition by Ang IV was dose dependent and was blocked by Divalinal. The fact that the Ang IV enhancement of normal LTP was blocked by losartan, an Ang II AT1 receptor antagonist, is puzzling since Divalinal had no effect on the inhibition of LTP by Ang II.

Angiotensin II blocks hippocampal long-term potentiation

We have found that injection of angiotensin II (AII) above the hippocampus in the intact rat blocks the induction of long-term potentiation (LTP) in perforant path-stimulated dentate granule cells. A minimum dose of 4.78 pmol AII was required for the complete blockade of LTP and this blockade was entirely prevented if the AII-specific antagonist saralasin was co-injected at a 50-fold molar excess. AII thus appears to act via AII receptors and does not cause non-specific inhibition. The injection of saralasin alone yielded LTP comparable to that obtained when vehicle was injected. Angiotensin III was found to be 40-50 fold less potent than AII in blocking LTP. Both AII and AII receptors of unknown function occur in the hippocampal formation. The results reported here suggest a role for these molecules in the control of hippocampal LTP.

Role of angiotensin II and AT1 receptors in hippocampal LTP

Results of a previous study showed that angiotensin II (AII) inhibited the induction of long-term potentiation (LTP) in hippocampal granule cells in response to dorsomedial perforant path stimulation in urethane-anesthetized rats. The results of present experiments demonstrate a dose-dependent inhibition of LTP induction under the same conditions due to ethanol (EtOH) administered by stomach tube and diazepam (DZ) injected IP. The inhibition of LTP induction by EtOH and DZ can be blocked by saralasin (SAR) applied directly to the dorsal hippocampus and by lorsartan (DuP 753) administered IP. Lorsartan or a metabolite crosses the blood-brain barrier because it also blocks the inhibition of LTP induction due to AII administration directly into the dorsal hippocampus. Lorsartan is a competitive antagonist of the AT1 subtype AII receptor. Therefore, the AII and the EtOH and DZ inhibition of LTP induction are mediated by the AII subtype receptor AT1. AIII and the AT2 antagonist PD123319 did not produce any significant effects. These in vivo effects can be reproduced in brain slices and therefore cannot be attributed to other factors, such as the urethane. In addition, electrical stimulation of the lateral hypothalamus (LH) inhibits LTP induction, and the inhibition can be blocked by SAR. These data on LH stimulation indicate that LH AII-containing neurons send axons into the hippocampus that inhibit the induction of LTP. These results not only provide new information on a neurotransmitter involved in the amnesic effects of benzodiazepines and ethanol-induced memory blackouts, but also testable hypotheses concerning recent observations that angiotensin converting enzyme (ACE) inhibitors elevate mood and improve certain cognitive processes in the elderly.