Juan Carlos Pineda

Caffeine and muscarinic antagonists act in synergy to inhibit haloperidol-induced catalepsy

The possible synergism between caffeine and muscarinic antagonists to inhibit haloperidol-induced catalepsy was investigated with the bar test in rats. Pretreatment with low doses of caffeine (1-3 mg/kg), a non-selective adenosine antagonist, dose dependently reduced the intensity and increased the onset latency of catalepsy induced by haloperidol (0.5-2 mg/kg). Similar effects were produced by the muscarinic antagonists atropine (4.1 mg/kg), and trihexyphenidyl (THP, 0.01-3 mg/kg). THP inhibited catalepsy intensity with an ED(50) of 0.38 mg/kg, and increased its onset latency with an ED(50) of 0.52 mg/kg. The anticataleptic effect of anticholinergics was potentiated when a low dose of caffeine (1 mg/kg) was applied simultaneously. In the presence of caffeine, THP inhibited catalepsy intensity with an ED(50) of 0.19 mg/kg, and prolonged the latency with an ED(50) of 0.30 mg/kg. The synergism was more evident when THP was administered at subthreshold doses that were unable to modify haloperidol-induced catalepsy when applied alone, but produced a clear inhibition of catalepsy when injected with caffeine. To assess whether repeated administration of caffeine could induce tolerance to the synergism with THP, a group of rats was pretreated with three daily doses of caffeine (1 mg/kg) for seven days, and the catalepsy test was performed on the eighth day. In these animals, caffeine was still able to enhance the anticataleptic actions of THP, suggesting that repeated administration of 1 mg/kg caffeine does not induce tolerance to the synergism with anticholinergics. These results indicate that low doses of caffeine enhance the anticataleptic actions of muscarinic antagonists, and leave open the possibility of using caffeine as adjunctive therapy to reduce the doses and the adverse effects of anticholinergics in Parkinsons disease.

5‐HT1A, 5‐HT2, and GABAB receptors interact to modulate neurotransmitter release probability in layer 2/3 somatosensory rat cortex as evaluated by the paired pulse protocol

Activation of γ‐aminobutyric acid B (GABAB) and 5‐hydroxytryptamine (5‐HT) receptors produces presynaptic inhibition at glutamatergic terminals in the rat neocortex. To evaluate interactions between these metabotropic receptors, field potentials were recorded in layer 2/3 of somatosensory cortex. In addition, the paired pulse (PP) protocol was used to measure changes in the ratio of the second/first extracellular synaptic potentials (S2/S1 ratio) as an index of glutamate release probability in the area. Lowering extracellular [Ca2+]o to 0.5 mM, increased the S2/S1 ratio by 318 ± 134%. 5‐HT (1 μM) increased the S2/S1 ratio by 61 ± 15%. In presence of the GABAA antagonist bicuculline (10 μM), 5‐HT increased the S2/S1 ratio by 98 ± 15%. This effect did not desensitize after two consecutive applications of the amine, and was dose dependent in the concentration range between 0.03–1 μM (EC50 = 2.36 × 10−7 mol/L). The increase of S2/S1 ratio induced by 5‐HT (1 μM) was blocked reversibly by the 5‐HT1A antagonist NAN‐190 (10–30 μM), but was unaffected by the selective GABAB antagonist CGP 52432 (1 μM). The action of 5‐HT was mimicked by the 5‐HT1A/7 agonist 8OH‐DPAT (10 μM), increasing the S2/S1 ratio by 84 ± 2%, a response that was unaffected by the 5‐HT2/7 antagonist ritanserin (2 μM). The 5‐HT1B agonist CP93129 (10 μM) had no effect. The GABAB agonist baclofen (1 μM) increased the S2/S1 ratio up to 308 ± 33%, which is similar to that produced by low [Ca2+]o. When the effect of baclofen was maximal, application of 5‐HT (1 μM) reversed the S2/S1 ratio back to 78 ± 27%, a result that was blocked by the 5‐HT2/7 antagonist ritanserin (100 nM). Notably, the interaction between the GABAB agonist and 5‐HT was order dependent, because enhancement of the S2/S1 ratio elicited by baclofen was not inhibited if 5‐HT was applied first. These results suggest a complex interaction between GABAB, 5‐HT1A, and 5‐HT2 receptors in layer 2/3 of rat somatosensory cortex. Activation of GABAB receptors induces PP facilitation (inhibits glutamate release) more efficiently than does activation of 5‐HT1A receptors. When the effect of GABAB receptor activation is maximal, however, the influence of 5‐HT changes to the opposite direction, inhibiting PP facilitation (increasing glutamate release) through activation of 5‐HT2 receptors.

Differential effects of caffeine on the antidepressant-like effect of amitriptyline in female rat subpopulations with low and high immobility in the forced swimming test.

The interaction of caffeine (1 mg/kg) and amitriptyline (15 mg/kg) on the immobility time (IT) during Porsolts forced swimming test (FST) was investigated in female Wistar rats. Akaikes Information Criterion indicated that the ITs recorded from 142 rats during the first day of the FST followed a bimodal distribution. Hence, the median (125.5 s) was used to classify the animals in subpopulations with low (<125.5 s, LI-rats) or high (>125.5 s, HI-rats) immobility. The paired t-test was used to compare the change of ITs between the first and second swimming sessions. Vehicle-treated animals had a significant increase of ITs during the second day of the test, either in LI-rats (77+/-12 s vs. 196+/-8 s, P<0.0001, n=6) or HI-rats (150+/-8 s vs. 201+/-10 s, P<0.02, n=6). In LI-rats amitriptyline only prevented the increase of ITs during the second session (74+/-27 s vs. 97+/-42 s, n=12), whereas in HI-rats the antidepressant produced a significant decrease of ITs during the second session (161+/-22 s vs. 118+/-32 s, n=7, P<0.02). While caffeine alone prevented the increase of ITs in both groups, the methylxanthine abolished the effect of amitriptyline in HI-rats (165+/-23 s vs. 165+/-46 s, n=9), leaving the antidepressant action unaffected in LI-rats (87+/-23 s vs. 96+/-58 s, n=9). These results suggest that the anti-immobility effect of amitriptyline in HI-rats is mediated in part by endogenous adenosine.

Locus Ceruleus Norepinephrine Release: A Central Regulator of CNS Spatio-Temporal Activation?

Norepinephrine (NE) is synthesized in the Locus Coeruleus (LC) of the brainstem, from where it is released by axonal varicosities throughout the brain via volume transmission. A wealth of data from clinics and from animal models indicates that this catecholamine coordinates the activity of the central nervous system (CNS) and of the whole organism by modulating cell function in a vast number of brain areas in a coordinated manner. The ubiquity of NE receptors, the daunting number of cerebral areas regulated by the catecholamine, as well as the variety of cellular effects and of their timescales have contributed so far to defeat the attempts to integrate central adrenergic function into a unitary and coherent framework. Since three main families of NE receptors are represented—in order of decreasing affinity for the catecholamine—by: α2 adrenoceptors (α2Rs, high affinity), α1 adrenoceptors (α1Rs, intermediate affinity), and β adrenoceptors (βRs, low affinity), on a pharmacological basis, and on the ground of recent studies on cellular and systemic central noradrenergic effects, we propose that an increase in LC tonic activity promotes the emergence of four global states covering the whole spectrum of brain activation: (1) sleep: virtual absence of NE, (2) quiet wake: activation of α2Rs, (3) active wake/physiological stress: activation of α2- and α1-Rs, (4) distress: activation of α2-, α1-, and β-Rs. We postulate that excess intensity and/or duration of states (3) and (4) may lead to maladaptive plasticity, causing—in turn—a variety of neuropsychiatric illnesses including depression, schizophrenic psychoses, anxiety disorders, and attention deficit. The interplay between tonic and phasic LC activity identified in the LC in relationship with behavioral response is of critical importance in defining the short- and long-term biological mechanisms associated with the basic states postulated for the CNS. While the model has the potential to explain a large number of experimental and clinical findings, a major challenge will be to adapt this hypothesis to integrate the role of other neurotransmitters released during stress in a centralized fashion, like serotonin, acetylcholine, and histamine, as well as those released in a non-centralized fashion, like purines and cytokines.

High-affinity inhibition of glutamate release from corticostriatal synapses by ω-agatoxin TK

Abstract To know which Ca2+ channel type is the most important for neurotransmitter release at corticostriatal synapses of the rat, we tested Ca2+ channel antagonists on the paired pulse ratio. ω-Agatoxin TK was the most effective Ca2+ channel antagonist (IC50=127 nM; maximal effect=211% (with >1 μM) and Hill coefficient=1.2), suggesting a single site of action and a Q-type channel profile. Corresponding parameters for Cd2+ were 13 μM, 178% and 1.2. The block of L-type Ca2+ channels had little impact on transmission, but we also tested facilitation of L-type Ca2+ channels. The L-type Ca2+ channel agonist, s-(−)-1,4 dihydro-2,6-dimethyl-5-nitro-4-[2-(trifluoromethyl)phenyl]-3-pyridine carboxylic acid methyl ester (Bay K 8644 (5 μM)), produced a 45% reduction of the paired pulse ratio, suggesting that even if L-type channels do not participate in the release process, they may participate in its modulation.

Clinical doses of citalopram or reboxetine differentially modulate passive and active behaviors of female Wistar rats with high or low immobility time in the forced swimming test.

The sensitivity of immobility time (IT) to antidepressant-drugs differs in rats expressing high or low motor activity during the forced swimming test (FST). However, whether this heterogeneity is expressed after the administration of the most selective serotonin and norepinephrine reuptake inhibitors (SSRIs and SNRIs, respectively) is unknown. We compared the influence of either the SSRI citalopram or the SNRI reboxetine with the tricyclic antidepressant amitriptyline on two subgroups of female Wistar rats expressing high IT (HI; at or above the mean value) or low IT (LI; below the mean) during the initial 5 min of the first session of the FST. None of the tested drugs increased motor activity in the open field test. When vehicle was applied to either HI or LI rats, IT increased in the second session of the FST. This increment concurred with a simultaneous climbing time (CT) decrement. When amitriptyline (15 mg/kg) was tested the CT increased for both HI and LI rats. This increment was accompanied by an IT decrement in HI and LI rats. Reboxetine (0.16 or 1 mg/kg) precluded IT and CT changes in both HI and LI rats and produced a swimming time reduction. Citalopram (0.4, 1, and 3 mg/kg) essentially mimicked the influence of reboxetine on the IT and CT in LI rats, as well as in HI rats, but in the latter case only at 3 mg/kg. Yet, at the dose of 10 mg/kg citalopram lacked this effect in both subgroups. No differences were detected when the IT of LI rats was evaluated with citalopram (3 mg/kg) during estrus or diestrus stage. These results show that clinical doses of citalopram produced an antidepressant-like effect selectively in LI rats, while amitriptyline or reboxetine produced this effect in both LI and HI animals.

Interlaminar differences of spike activation threshold in the auditory cortex of the rat

The neural circuits of the auditory cortex are a substrate for the dual purpose of representing and storing the auditory signal on one hand, and sending its relevant features to other cortical and subcortical areas on the other hand. The ability to process and transform the signal crucially depends on achievement of the neuronal spike threshold following spatiotemporal summation of the synaptic signals. We used patch-clamp recording in a thin slice preparation to compare neuronal responses to current injection of layer II/III and layer V neurons. We found that while the two classes of neurons do not differ in passive neuronal properties, layer II/III neurons possess a lower firing threshold relative to layer V neurons (-44.8 +/- 2.4 mV vs. -34.3 +/- 4.0 mV). We speculate that a lower spiking threshold in layer II/III neurons might favor local intracolumnar activation for representation and storage of the auditory information whereas a more positive spiking threshold for layer V neurons may prevent unnecessary cortical spread of a scarcely processed signal.

δ Opioids reduce the neurotransmitter release probability by enhancing transient (KV4) K+-currents in corticostriatal synapses as evaluated by the paired pulse protocol

Field recordings were used to determine the influence of delta-opioid receptor activation on corticostriatal synaptic transmission. Application of the selective delta-opioid receptor agonist, [Tyr-D-Pen-Gly-Phe-D-Pen]-enkephalin (DPDPE, 1 microM), decreased the amplitude of the field-excitatory synaptic potential and at the same time increased the paired pulse ratio (PPR) suggesting a presynaptic site of action. This response reversed rapidly when DPDPE was washed and blocked by 1 nM of the selective delta-receptor antagonist naltrindole. Neither omega-conotoxin GVIA (1 microM) nor omega-agatoxin TK (400 nM), blockers of N- and P/Q-type Ca2+-channels, respectively, nor TEA (1 mM), blocker of some classes of K+-channels, occluded the effects of DPDPE. Instead, 1 mM 4-AP or 400 microM Ba2+ occluded completely the effects of DPDPE. Therefore, the results suggest that the modulation by delta opioids at corticostriatal terminals is mediated by transient (KV4) K+-conductances.

Long-lasting resistance to haloperidol-induced catalepsy in male rats chronically treated with caffeine.

Chronic caffeine consumption has been inversely associated with the risk of developing Parkinsons disease. Here we assessed whether chronic caffeine treatment increases the resistance of male Wistar rats to haloperidol (1mg/kg, s.c.)-induced catalepsy, measured in the bar test at 15 min intervals during 3h. Caffeine (5mg/kg/day) was delivered for 6 months via drinking water. Control rats received only tap water. Treatments began when animals were 3-4 months old. In order to unveil long-lasting catalepsy refractoriness not attributable to the presence of caffeine in the brains of rats, they were evaluated from day 18 to day 27 after caffeine withdrawal, a time that is far in excess for the full excretion of a caffeine dose in this species. The average cataleptic immobility measured in caffeine-treated rats (n=23) was 1148+/-140 s, a value 34+/-8% lower than that recorded in control animals (n=20), whose mean immobility was 1736+/-137 s (P=0.0026, t-test). The percentage of catalepsy reduction measured in caffeine-treated rats evaluated on days 18-20 after caffeine discontinuation (-32+/-13%, n=12, P<0.05) was comparable to the catalepsy decrease recorded in those animals tested on days 21-27 (-36+/-10%, n=11, P<0.02), a finding compatible with the notion that the effect was indeed mediated by enduring changes of brain functioning and not by the physical presence of caffeine or its metabolites. Caffeine-treated rats also had higher catalepsy latency scores compared with control rats (P<0.01, U-test). The present findings show that chronic consumption of caffeine produces perdurable resistance to catalepsy induced by dopamine receptor blockade, possibly through enhancement of dopamine transmission, giving further support to the epidemiological results indicating that prolonged caffeine consumption affords neuroprotection against Parkinsons disease.

N‐type calcium channels mediate a GABAB presynaptic modulation in the corticostriatal synapse in turtle's paleostriatum augmentatum

Spikes population evoked by a paired pulse protocol were used to assess the influence of GABAA and GABAB receptors agonists and antagonists on the synaptic potentials and in the S2/S1 ratio in a paired pulse (PP) protocol in the cortico‐paleostriatum augmentatum synapses of the turtle. GABAA agonist, muscimol, decreased the amplitude of synaptic responses whereas the facilitation produced with the PP protocol did not change, suggesting a postsynaptic action for GABAA receptors. GABAB agonist, baclofen, enhanced paired pulse ratio indicating a presynaptic modulation through the GABAB receptor. Selective antagonists for N‐ and P/Q‐type Ca2+‐channels also enhanced paired pulse ratio, suggesting that any of these channel types may be involved in neurotransmitter release. However, the strong paired pulse facilitation produced by baclofen was occluded by blocking the N‐type Ca2+ channels with ω‐conotoxin GVIA (1 μM), but not by the blockage of P/Q‐type Ca2+ channels with ω‐agatoxin TK (400 nM). These data suggest that N and P/Q channels participate in the neurotransmitter release, whereas only N‐type Ca2+ channels are involved in the presynaptic modulation of GABAB in the corticostriatal synapse of the turtle. Synapse 63:855–862, 2009.