C. Infante

Copper suppresses hippocampus LTP in the rat, but does not alter learning or memory in the morris water maze.

The objective of our study was to determinate the effect of copper on long-term potentiation (LTP) in hippocampus slices and a learning test in the Morris Water Maze (MWM). A group of adult Wistar rats received intraperitoneal (ip) injections of 1 mg/kg of CuSO(4) dissolved in saline for 30 consecutive days (Cu.R). A group of control rats (Sal.R), received saline by the same routes and duration. After this period, every individual of both groups was submitted to learning in MWM. Once the learning was completed, the LTP was studied in slices of hippocampus of both groups. The statistical assessment shows that the rats in both groups did not show significant differences in their progressive learning, notwithstanding that group Cu.R had 14.2 times more copper in their hippocampus and 16.7 times more in the visual cortices than in those of group Sal.R. On the other hand, the neurons of CA1 in hippocampus slices of Sal.R showed a significant development of LTP, but this was not observed in group Cu.R. In a second situation, 13 rats received training in MWM. Then, a group of 6 animals were injected with copper i.p. at the dose and time previously described. The 7 other animals were administered saline. Afterward, both groups were retrained in the MWM. The results obtained in Cu.R were similar to those obtained in Sal.R. Both groups maintained the concentrations of copper in the hippocampus indicated above, nonetheless, only the hippocampus slices of Cu.R did not show LTP. The spatial learning behavior of the rats was not affected by high copper concentration.

Involvement of spinal cord BDNF in the generation and maintenance of chronic neuropathic pain in rats

Brain-derived neurotrophic factor (BDNF) is involved in neuronal survival and synaptic plasticity of the central and peripheral nervous system. In chronic pain, plastic changes in dorsal horn neurons contribute to a phenomenon of hypersensitivity to pain sensation that is maintained over time, known as central sensitization. This process is accompanied by BDNF overexpression, but the role of BDNF in the generation and maintenance of the hyperalgesic phenomenon is still unclear. The present study was aimed to investigate if exogenous BDNF administered to the rat spinal cord, in addition to trigger pain, participates in the maintenance of the central sensitization process (i.e., pain persistence) and to determine if the pain generated is comparable to that observed in a neuropathic pain model. Results showed that a single intrathecal injection of 0.003 ng of BDNF was able to decrease the nociceptive threshold (Randall-Selitto test) in normal rats, for at least a 42-day period. Furthermore, the hyperalgesia generated was comparable to that observed in rats with a 42-day history of mononeuropathy. Increasing the dose or administering additional doses of BDNF resulted neither in additional effectiveness in reducing the pain threshold nor in the prolongation of the hyperalgesic effect, thus showing that central sensitization induced by BDNF is a dose-independent, all-or-none process. It is concluded that BDNF alone is sufficient for generating a long-lasting neural excitability change in the spinal cord via tyrosine kinase B receptor signaling, similar to that observed in chronic pain models such as neuropathy.

Interference of chronically ingested copper in long-term potentiation (LTP) of rat hippocampus

The objective of our study was to find the evidence of copper interaction in LTP, motivated by copper involvement in neurodegenerative illness, like Parkinson, Alzheimer and Amyotrophic Lateral Sclerosis, and we initiated the study of this element in the LTP. For this purpose we used hippocampus slices of rats chronically consuming copper dissolved in water (CuDR; n=26) and non-copper-consuming rats (CR; n=20). The CuDR rats received 8--10 mg/day during 20--25 days. Electrophysiological tests showed absence of LTP in CuDR slices, contrary to CR slices. The stimulus-response test applied before and after LTP showed significant increases of synaptic potential in the CR group. This did not occur in the CuDR group, except for the initial values, which probably seem associated to an early action of copper. The paired-pulse (PP) test, applied to CR and CuDR prior to tetanic stimulation, showed a significant reduction in PP, for the 20-, 30- and 50-ms intervals in CuDR. At the end of the experiments, copper concentration was 54.2 times higher in CuDR slices, compared to the concentration present in CR slices. Our results show that copper reduces synaptic sensibility and also the facilitation capability. These effects represent a significant disturbance in the plasticity phenomenon associated with learning and memory.

Simultaneous unitary neuronal activity in both superior colliculi and its relation to eye movements in the cat

The analysis of simultaneous unitary neuronal activity related to eye movement and recorded in both superior colliculi has shown a mirror-functioning image. Increase of the frequency discharge in a collicular unit, was associated with a decrease of the frequency discharge in the contralateral superior colliculus unit. This unitary neuronal reciprocal behaviour was observed each time a horizontal or oblique eye movement was produced. It is possible that this reciprocal functioning between neurones in both superior colliculi could exert an important influence on oculomotor brainstem structures. These results give a better idea of the role played by both superior colliculi in the control of conjugate eye movements.

Expression of nitric oxide synthase isoforms in the dorsal horn of monoarthritic rats: effects of competitive and uncompetitive N-methyl-D-aspartate antagonists

Chronic pain is associated with N-methyl-D-aspartate (NMDA) receptor activation and downstream production of nitric oxide, which has a pivotal role in multisynaptic local circuit nociceptive processing in the spinal cord. The formation of nitric oxide is catalyzed by three major nitric oxide synthase (NOS) isoforms (neuronal, nNOS; inducible, iNOS; endothelial, eNOS), which are increased in the spinal cord of rodents subjected to some tonic and chronic forms of experimental pain. Despite the important role of NOS in spinal cord nociceptive transmission, there have been no studies exploring the effect of NMDA receptor blockade on NOS expression in the dorsal horn during chronic pain. Furthermore, NOS isoforms have not been fully characterized in the dorsal horn of animals subjected to arthritic pain. The aim of this work was therefore to study the expression of nNOS, iNOS and eNOS in the dorsal horns of monoarthritic rats, and the modifications in NOS expression induced by pharmacological blockade of spinal cord NMDA receptors. Monoarthritis was produced by intra-articular injection of complete Freunds adjuvant into the right tibio-tarsal joint. At week 4, monoarthritic rats were given either the competitive NMDA antagonist (±)-3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP) or the uncompetitive NMDA antagonist ketamine. After 6 and 24 hours, animals were killed and posterior quadrants of the lumbar spinal cord were dissected. Sample tissues were homogenized and subjected to immunoblotting with anti-nNOS, anti-iNOS or anti-eNOS monoclonal antibodies. The nNOS isoform, but not the iNOS and eNOS isoforms, were detected in the dorsal horns of control rats. Monoarthritis increased the expression of nNOS, iNOS and eNOS in the dorsal horns ipsilateral and contralateral to the inflamed hindpaw. Intrathecal administration of CPP and ketamine reduced nNOS expression in monoarthritic rats but increased the expression of iNOS and eNOS. Results suggest that blockade of spinal cord NMDA receptors produces complex regulatory changes in the expression of NOS isoforms in monoarthritic rats that may be relevant for nitridergic neuronal/glial mechanisms involved in the pathophysiology of monoarthritis and in the pharmacological response to drugs interacting with NMDA receptors.

Antinociceptive effect and interaction of uncompetitive and competitive NMDA receptor antagonists upon capsaicin and paw pressure testing in normal and monoarthritic rats

&NA; We assessed whether intrathecal administration of the uncompetitive and competitive NMDA receptor antagonists ketamine and (±)CPP, respectively, could produce differential modulation on chemical and mechanical nociception in normal and monoarthritic rats. In addition, the antinociceptive interaction of ketamine and (±)CPP on monoarthritic pain was also studied using isobolographic analysis. Monoarthritis was produced by intra‐articular injection of complete Freund’s adjuvant into the tibio‐tarsal joint. Four weeks later, the antinociceptive effect of intrathecal administration of the drugs alone or combined was evaluated by using the intraplantar capsaicin and the paw pressure tests. Ketamine (0.1, 1, 10, 30, 100, 300 and 1000 μg i.t.) and (±)CPP (0.125, 2.5, 7.5, 12.5, 25 and 50 μg i.t.) produced significantly greater dose‐dependent antinociception in the capsaicin than in the paw pressure test. Irrespective of the nociceptive test employed, both antagonists showed greater antinociceptive activity in monoarthritic than in healthy rats. Combinations produced synergy of a supra‐additive nature in the capsaicin test, but only additive antinociception in paw pressure testing. The efficacy of the drugs, alone or combined, is likely to depend on the differential sensitivity of tonic versus phasic pain and/or chemical versus mechanical pain to NMDA antagonists.

Rotational behavior in the cat induced by electrical stimulation of the pulvinar-lateralis posterior nucleus complex: role of the cholinergic system.

We studied the involvement of the cholinergic system in the contralateral head-eye-body turning induced in the cat through stimulation of the pulvinar-lateralis posterior nucleus complex (P-LP). In 17 cats through a cannula aimed at the P-LP, agonists and antagonists of the cholinergic system were injected. The electrical activity of the P-LP could be recorded through the same cannula or through electrodes attached to it. In addition, electrodes were implanted ipsilaterally in the dorsal hippocampus, caudate nucleus, amygdala, and superior colliculus to record through them and through one screw placed on the skull the electrical activity of those structures and of the cortical P-LP projection. Seven days after surgery, carbachol, an agonist of the cholinergic system was injected in the P-LP, and the behavior and electrical activity of the unrestrained cat (previously accustomed to a plastic cage) were recorded. A control volume of 0.9% NaCl was always injected previously. The usual drug volume injected was 1 microliter; occasionally, 2 microliter were injected. Weekly or biweekly sessions were conducted to determine (a) the threshold for cholinergic activation, (b) the threshold for turning behavior, (c) the blocking effect of local atropine sulfate injected previously, (d) the effect of haloperidol previously injected (locally or systemically), and (e) the effect of dioxolane, an exclusive muscarinic agonist. In 14 of 17 cats, contralateral turning behavior was evoked by carbachol. In two of the three cats that did not respond to carbachol, dioxolane induced turning. The effect of dioxolane was similar to that of carbachol when tried in five cats. Besides turning behavior, carbachol produced numerous symptoms due to cholinergic activation. Atropine blocked the rotational effect of carbachol in all cats, and haloperidol blocked it in 68% of them. Electrolytic coagulation of the dorsal hippocampus surrounding the P-LP did not disturb the effects induced by carbachol. These experiments show that both systems of the P-LP, cholinergic and catecholaminergic, are involved in the contralateral turning. We conclude that the effect induced by carbachol is due to activation of muscarinic receptors because it is totally blocked by local atropine sulfate and is reproduced by dioxolane, an exclusive muscarinic agonist.

Output pathway for turning behavior from the neostriatum and substantia nigra in cats.

The goal of the present work was to study the output pathway of the information for turning behavior originating in the striatum and coursing through the substantia nigra pars reticulata (SNR). In 45 adult cats distributed in 3 groups, Ni-Cr electrodes were implanted in the caudate nucleus and substantia nigra pars reticulata and depending on the animal group in the superior colliculus (SC), nucleus ventralis lateralis/nucleus ventralis medialis (VL/VM) complex or nucleus tegmenti pedunculopontinus (TPP) of one cerebral hemisphere. The threshold current required to evoke turning behavior was determined in each animal for each implantation site. An electrolytic lesion of the superior colliculus, the VL/VM complex or the nucleus tegmenti pedunculopontinus was carried out in each group of cats. The effects of the lesions on behavior and on the electrical threshold currents were determined and compared with the prelesion values. Finally the extent of the lesions and electrode positions were analyzed. The results show that the substantia nigra pars reticulata is the structure with the lowest thresholds for turning and that the superior colliculus appears to be more relevant for carrying the information for turning than either the VL/VM complex or the nucleus tegmenti pedunculopontinus.

Correlation between pulvinar-lateralis posterior complex unit activity and eye movements in the cat

In 15 encéphale isolé cats, 130 units were recorded in the pulvinar-lateralis posterior complex (P-LP); 19 units responded in relation to horizontal eye movements (15%), 9 of which discharged when ipsiversive movements were recorded and the remaining 10 to contraversive movements. Three units discharged during spontaneous nystagnoid eye movement. All units always responded after initiation of the eye movement, with a latency range between 50 and 250 ms. The eye movement-related units were preferentially located (80%) in the border between the pulvinar and the lateralis posterior complex. Our results show the presence of eye movement-related units in the cats P-LP, and their probable participation in ocular motility.

Study of the morphological, electrophysiological and behavioral effects of unilateral kainic acid injection into the cat's substantia nigra.

Morphological, electrophysiological and behavioral studies were carried out in cats after unilateral kainic acid injection in the substantia nigra. A forced contralateral head turning and compulsive circling was observed after surgery. Fifteen days after, when asymmetry disappeared, apomorphine induced an ipsilateral head and body turning, that was blocked by haloperidol. The percentage of turning, after electrical stimulation in the superior colliculus or pulvinar-lateralis posterior complex, was affected by substantia nigra lesion. This work demonstrates that the nigro-pulvinar-lateral posterior and the nigrotectal projection modulate the capability of electrical stimulation of the target structures to elicit turning, and after unilateral substantia nigra lesion, two opposite directions of asymmetry appear, which are time-dependent and modulated by different neurotransmitters.