We identified a new form of activity-dependent modulation of the afterhyperpolarization (AHP) in tactile (T) sensory neurons of the leech Hirudo medicinalis. Repetitive intracellular stimulation with 30 trains of depolarizing impulses at 15-s inter-stimulus interval (ISI) led to an increase of the AHP amplitude (~60% of the control). The enhancement of AHP lasted for >/=15 min. The AHP increase was also elicited when a T neuron was activated by repetitive stimulation of its receptive field. The ISI was a critical parameter for the induction and maintenance of AHP enhancement. ISI duration had to fit within a time window with the upper limit of 20 s to make the training effective to induce an enhancement of the AHP amplitude. After recovery from potentiation, AHP amplitude could be enhanced once again by delivering another training session. The increase of AHP amplitude persisted in high Mg(2+) saline, suggesting an intrinsic cellular mechanism for its induction. Previous investigations reported that AHP of leech T neurons was mainly due to the activity of the Na(+)/K(+) ATPase and to a Ca(2+)-dependent K(+) current (I(K/Ca)). In addition, it has been demonstrated that serotonin (5HT) reduces AHP amplitude through the inhibition of the Na(+)/K(+) ATPase. By blocking the I(K/Ca) with pharmacological agents, such as cadmium and apamin, we still observed an increase of the AHP amplitude after repetitive stimulation, whereas 5HT application completely inhibited the AHP increment. These data indicate that the Na(+)/K(+) ATPase is involved in the induction and maintenance of the AHP increase after repetitive stimulation. Moreover, the AHP increase was affected by the level of serotonin in the CNS. Finally, the increase of the AHP amplitude produced a lasting depression of the synaptic connection between two T neurons, suggesting that this activity-dependent phenomenon might be involved in short-term plasticity associated with learning processes.

Increasing evidence indicates that modulation of Na(+)/K(+) ATPase activity is involved in forms of neuronal and synaptic plasticity. In tactile (T) neurons of the leech Hirudo medicinalis, Na(+)/K(+) ATPase is the main determinant of the afterhyperpolarization (AHP), which characterizes the firing of these mechanosensory neurons. Previously, it has been reported that cAMP (3',5'-cyclic adenosine monophosphate), which mediates the effects of serotonin (5HT) in some forms of learning in the leech, negatively modulates Na(+)/K(+) ATPase activity, thereby reducing the AHP amplitude in T neurons. Here, we show that a transient inhibition of Na(+)/K(+) ATPase can affect the synaptic connection between two ipsilateral T neurons. Bath application of 10 nm dihydroouabain (DHO), an ouabain analogue, causes an increase in the amplitude of the synaptic potential (SP) recorded in the postsynaptic element when a test stimulus is applied in the presynaptic neuron. Iontophoretic injection of cAMP into the presynaptic T neuron also produces an increase of SP. Simulations carried out by using a computational model of the T neuron suggest that a reduction of the pump rate and a consequent depression of the AHP might facilitate the conduction of action potentials to the synaptic terminals. Moreover, nearly intact leeches injected with 10 nm DHO respond with a swimming episode more quickly to an electrical stimulation, which selectively activates T neurons exhibiting sensitization of swimming induction. Collectively, our results show that inhibition of Na(+)/K(+) ATPase is critical for short-term plasticity.

Pituitary adenylate cyclase-activating polypeptide (PACAP-38) is a member of the vasointestinal polypeptide (VIP)/secretin/glucagon family of neuropeptides for which neuroregulatory functions have been postulated. PACAP-38 receptors are expressed in different brain regions, including hippocampus. In this study, we examined the dose-dependent effects of PACAP-38 on the excitatory postsynaptic field potential (fEPSP) evoked at the Schaffer collateral-CA1 synapse in rat hippocampal slices. Bath application of low dose (0.05 nM) of PACAP-38 induced long-lasting facilitation of the fEPSP. This enhancement was blocked by the cholinergic receptor antagonist atropine and partially by the NMDA receptor antagonist 2-amino-5-phosphonovalerate (APV) and therefore, shares a common mechanism with LTP. In contrast, a high dose (1 µM) of PACAP-38 induced a persistent depression of the fEPSP that was not blocked by antagonists of cholinergic receptors (i.e., atropine and mecamylamine), adenosine receptors (i.e., DCPCX), or glutamatergic NMDA receptors (APV). Intermediate doses (0.1-0.5 µM) of PACAP-38 produced an initial decrease of the fEPSP followed by an enhancement. This decrease was not blocked by atropine whereas the facilitation was. These results show that PACAP-38 modulates CA1 synaptic transmission in a dose-dependent manner and that the peptide interacts with cholinergic and glutamatergic systems.Pituitary adenylate cyclase-activating polypeptide (PACAP-38) is a 38-amino acid peptide that was first isolated from ovine hypothalamus for its ability to stimulate adenylyl cyclase in rat anterior pituitary cells (Arimura 1992). PACAP-38 exhibits high sequence identity with vasoactive intestinal peptide (VIP), distinguishing PACAP-38 as a member of the VIP-secretin-glucagon family of peptides. The aminoacid sequence of PACAP-38 has been remarkably conserved during evolution, suggesting that PACAP-38 regulates important physiological functions (Arimura 1992;Masuo et al. 1993). Two receptors for PACAP-38 have been identified: type I receptors, which are positively coupled to adenylyl cyclase and phospholipase C, and type II receptors, which have only been linked to adenylyl cyclase (Spengler et al. 1993). PACAP-38 receptors are mainly distributed in the central nervous system including the hippocampus (Masuo et al. 1992(Masuo et al. , 1993.PACAP-38 modulates synaptic activity in several neuronal regions. For example, PACAP-38 enhances in a dosedependent manner the spontaneous release of acetylcholine (ACh) from septal cholinergic fibers in the dorsal hippocampus (Masuo et al. 1993). An excitatory action of PACAP-38 on glutamatergic N-methyl-D-aspartate (NMDA) receptors has also been reported in cortical neurons (Martin et al. 1995;Stella and Magistretti 1996;Liu and Madsen 1997) and in sympathetic preganglionic neurons of neonatal rat (Lai et al. 1997;Wu and Dun 1997). In addition, a high concentration of PACAP-38 (1-3 µM) induces a long-lasting depression of the field excitatory postsynaptic potential (fEPSP) at hippocampal ...

In the present study we have extended our previous findings about the effects of 10 minutes of passive mandibular extension in anesthetized Wistar rats. By prolonging the observation time to 3 hours, we showed that 10 minutes mandibular extension caused a significant reduction of the mean arterial blood pressure and heart rate respect to baseline values, which persisted up to 160 minutes after mandibular extension. These effects were accompanied by a characteristic biphasic response of pial arterioles: during mandibular extension, pial arterioles constricted and after mandibular extension dilated for the whole observation period. Interestingly, the administration of the opioid receptor antagonist naloxone abolished the vasoconstriction observed during mandibular extension, while the administration of Nω-Nitro-L-arginine methyl ester, a nitric oxide synthase inhibitor, abolished the vasodilation observed after mandibular extension. Either drug did not affect the reduction of mean arterial blood pressure and heart rate induced by mandibular extension. By qRT-PCR, we also showed that neuronal nitric oxide synthase gene expression was significantly increased compared with baseline conditions during and after mandibular extension and endothelial nitric oxide synthase gene expression markedly increased at 2 hours after mandibular extension. Finally, western blotting detected a significant increase in neuronal and endothelial nitric oxide synthase protein expression. In conclusion mandibular extension caused complex effects on pial microcirculation involving opioid receptor activation and nitric oxide release by both neurons and endothelial vascular cells at different times.

Previous data have shown both in the rat and in the human that a single mandibular extension lasting 10 min induces a significant important and prolonged reduction in blood pressure and heart rate, affecting also rat pial microcirculation by the release of endothelial factors. In the present work, we assessed whether repeated mandibular extension could further prolong these effects. We performed two mandibular extensions, the second mandibular extension being applied 10 min after the first one. The second mandibular extension produced a reduction in blood pressure and heart rate for at least 240 min. As in the case of a single mandibular extension, pial arterioles dilated persisting up to 140 min after the second extension. Spectral analysis on 30 min recordings under baseline conditions and after repetitive mandibular extensions showed that the pial arterioles dilation was associated with rhythmic diameter changes sustained by an increase in the frequency components related to endothelial, neurogenic, and myogenic activity while a single mandibular extension caused, conversely, an increase only in the endothelial activity. In conclusion, repetitive mandibular extension prolonged the effects of a single mandibular extension on blood pressure, heart rate and vasodilation and induced a modulation of different frequency components responsible of the pial arteriolar tone, in particular increasing the endothelial activity.

No abstract

A large variety of distinct locally connected GABAergic cells are present in the hippocampus. By releasing GABA into principal cells and interneurons, they exert a powerful control on neuronal excitability and are responsible for network oscillations crucial for information processing in the brain. Here, whole-cell patch clamp recordings in current and voltage clamp mode were used to study the functional role of nicotinic acetylcholine receptors (nAChRs) on the firing properties of stratum oriens interneurons in hippocampal slices from transgenic mice expressing enhanced green fluorescent protein in a subpopulation of GABAergic cells containing somatostatin (GIN mice). Unexpectedly, activation of nAChRs by nicotine or endogenously released acetylcholine strongly enhanced spike frequency adaptation. This effect was blocked by apamin, suggesting the involvement of small calcium-dependent potassium channels (SK channels). Nicotine-induced reduction in firing frequency was dependent on intracellular calcium rise through calcium-permeable nAChRs and voltage-dependent calcium channels activated by the depolarizing action of nicotine. Calcium imaging experiments directly showed that nicotine effects on firing rate were correlated with large increases in intracellular calcium. Furthermore, blocking ryanodine receptors with ryanodine or sarcoplasmic-endoplasmic reticulum calcium ATPase with thapsygargin or cyclopiazonic acid fully prevented the effects of nicotine, suggesting that mobilization of calcium from the internal stores contributed to the observed effects. By regulating cell firing, cholinergic signalling through nAChRs would be instrumental for fine-tuning the output of stratum oriens interneurons and correlated activity at the network level.

Taurisolo is a pomace extract from Aglianico Grapes, a wine cultivar native to Campania (Southern Italy). It exhibits a very high polyphenolic content and, consumed as a nutraceutical, is effective in reducing the level of Trimethylamine N-oxide (TMAO), a cardiovascular disease risk factor marker. We here show the effects of Taurisolo on rat brain microvascular alterations induced by a diminution in cerebral blood flow (CBFD) for 30 min, due to bilateral common carotid artery occlusion, and subsequent blood flow restoration (CBFR) for 60 min. The rat pial microcirculation was investigated by intravital fluorescence microscopy through a parietal closed window implanted into the skull bone. The rat pial arterioles were classified according to Strahler's ordering scheme, from smaller penetrating arterioles up to the larger ones. Western blotting analysis and mass spectrometry (MS)-based metabolomics were used to investigate the expression of endothelial nitric oxide synthase (eNOS) or the presence of peroxidized cardiolipin and several inflammatory mediators, respectively. Radical Oxygen Species (ROS) formation and neuronal loss were assessed. In rats CBFD and CBFR caused a decrease in arteriolar diameter, increase in fluorescent leakage and in adhesion of leukocytes to venular walls, reduction in the length of perfused capillaries and increment of ROS formation with large infarct size. Taurisolo , intravenously or orally administered, induced pial arteriolar dilation (up to >30% of baseline), prevented fluorescent leakage, adhesion of leukocytes, ROS formation, while facilitated capillary perfusion and significantly reduced infarct size. These effects were accompanied by an increase in eNOS expression. Mass-spectrometry metabolomics analysis detected a marked decrease in the amount