Masashi Sasa

Excitatory and inhibitory effects of dopamine on neuronal activity of the caudate nucleus neurons in vitro

Effects of dopamine on the rat caudate nucleus neurons were examined in a slice preparation using an intracellular recording technique. Perfusion of the bath with a low concentration (1 microM) of dopamine produced a depolarization concomitant with an increase in the spontaneous firing and the number of action potentials evoked by a depolarizing pulse applied into the cells. In contrast, higher concentrations (100-500 microM) of dopamine inhibited the spontaneous and current-induced firings without apparent effects on the resting membrane potential. In addition, during application of a high concentration (100 microM) of dopamine there was a marked elevation of the threshold potential of the action potential elicited by a higher depolarizing current. Simultaneous application of haloperidol (0.5-5 microM) antagonized both excitatory and inhibitory effects induced by the low and high concentrations of dopamine, respectively. In addition, the excitatory effect induced by a low concentration (1 microM) of dopamine was antagonized by domperidone (0.5 microM), a selective D2 receptor antagonist, while the inhibitory effect by a high concentration (100 microM) was blocked by SCH 23390, a selective D1 receptor antagonist. These results strongly suggest that the postsynaptic sites of caudate nucleus neurons have at least two subtypes of dopamine receptors (D1 and D2 receptors) that mediate inhibitory and excitatory responses of the neuron to dopamine, respectively.

The dopamine D1 receptor agonist SKF 38393 suppresses detrusor hyperreflexia in the monkey with parkinsonism induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP).

A pharmacological study using monkeys, in which parkinsonism was induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), was undertaken to elucidate the mechanism underlying urinary bladder dysfunctions in Parkinsons disease. Under ketamine anesthesia, cystometrograms showed that, in MPTP-treated monkeys, a contraction of the urinary bladder was induced with smaller bladder volume than that in normal monkeys. In MPTP-treated monkeys, subcutaneously injected SKF 38393, a dopamine D1 receptor agonist, significantly increased the bladder volume and pressure thresholds for inducing the micturition reflex without affecting those in normal monkeys. In contrast, subcutaneous injections of quinpirole, a dopamine D2 receptor agonist, and apomorphine, a dopamine D1 and D2 receptor agonist, slightly, but significantly reduced the volume threshold of the bladder for the micturition reflex in both normal and MPTP-treated groups. These results indicate that, in parkinsonism, the degeneration of dopaminergic neurons in the substantia nigra leads to the detrusor hyperreflexia, probably due to a failure of activation of dopamine D1 receptors.

Noradrenaline-mediated inhibition by locus coeruleus of spinal trigeminal neurons

Abstract Inhibitory effects of conditioning stimulation of the locus coeruleus (LC) on the neuron activity in spinal trigeminal nucleus (STN) were investigated in gallamine-immobilized cats. Field potentials of STN and spike potentials of single relay neurons in STN were orthodromically elicited by ipsilateral alveolar nerve stimulation and antidromically by stimulation of contralateral medial lemniscus. Conditioning stimuli were applied to LC and sensory cortex (SC) at various C-T intervals. In tracking experiments near the LC region, conditioning stimulation of LC itself produced the most pronounced decrease in amplitude of the STN field potentials. Orthodromic spikes of STN single neurons were significantly reduced by conditioning stimulation of LC as well as SC. In reserpine-treated animals, however, conditioning stimulation of LC failed to produce a decrease in the number of orthodromic spikes, while the inhibitory effect of SC conditioning stimulation remained unaffected. Under these circumstances, intravenous L -dopa and intraventricular noradrenaline reproduced an inhibitory effect of LC conditioning stimulation on orthodromic spike generation, while such an effect was not seen with either dopamine or serotonin. Antidromic spike was unaltered by any of these treatments. Histochemically, catecholamine fluorescence in LC was entirely eliminated after reserpine-treatment, but was restored after L -dopa injection. These results strongly suggest that noradrenaline released from the terminals of neurons originating in LC produces an inhibition of transmission in the STN relay neurons.

Coexistence of inhibitory dopamine D-1 and excitatory D-2 receptors on the same caudate nucleus neurons

Microiontophoretic studies using cats anesthetized with alpha-chloralose were performed to determine whether or not dopamine D-1 and D-2 receptors co-exist in the same caudate nucleus (CN) neurons that receive inputs from the substantia nigra (SN), and in which spikes elicited by SN stimulation were blocked by domperidone, a selective D-2 antagonist. Iontophoretic application of dopamine produced a dose-dependent inhibition of spontaneous firing in 2 of 4 spontaneously active CN neurons and an increase in firing in the remaining 2 neurons. However, dopamine inhibited the glutamate-induced firing in 31 of 32 CN neurons that were not spontaneously active. Similar inhibition with iontophoretically applied SKF 38393, a selective D-1 agonist, was observed in 33 of 34 spontaneously inactive neurons tested. When the effects of dopamine, SKF 38393 and bromocriptine (D-2 agonist) were examined on the same CN neurons, the inhibitory effects of both dopamine and SKF 38393 were seen in 14 of 15 neurons, and both an inhibition by SKF 38393 and an excitation by bromocriptine were observed in 15 of 17 neurons. The inhibitory effects of dopamine and SKF 38393 were antagonized by haloperidol and SCH 23390 (D-1 antagonist) without being affected by domperidone. Furthermore, the dopamine-induced inhibition was converted to an excitation during simultaneous application of SCH 23390 in 6 of 10 CN neurons, and this excitation was antagonized by domperidone. These results strongly suggest that the inhibitory D-1 and excitatory D-2 receptors co-exist on the same CN neurons receiving inputs from the SN.

Attenuation of morphine analgesia in rats with lesions of the locus coeruleus and dorsal raphe nucleus

The nociceptive reflex activity and analgesic effect of morphine were studied in rats using the hind paw stimulation test. The stimulation threshold was significantly increased in animals with bilateral destruction of the locus coeruleus (LC), and was reduced after lesion of the dorsal raphe nucleus (DR). LC lesions produced a selective lowering of noradrenaline (NA) content in the forebrain, while DR lesions resulted in a reduction in serotonin levels. Lesioning both LC and DR significantly reduced both NA and serotonin contents even when the stimulation threshold was not altered. Morphine produced a significant and dose-dependent elevation of the stimulation threshold in sham-operated animals, while morphine analgesia was almost completely inhibited by destruction of LC, DR and both the nuclei. These results imply that a depression of LC-mediated noradrenergic tone results in a decreased sensitivity to painful stimuli, whereas a reduction of raphe-derived serotonergic tone produces the opposite effect against LC. It is suggested, however, that both of these monoamines from the LC and DR are necessary for the analgesic effect of morphine.

Attenuation by PACAP of glutamate-induced neurotoxicity in cultured retinal neurons

The effects of pituitary adenylate cyclase activating polypeptides (PACAPs: PACAP27, PACAP38) on glutamate-induced neurotoxicity were examined using cultured retinal neurons obtained from 3- to 5-day old Wistar rats. Cell viability was evaluated by double staining with fluorescein diacetate and propidium iodide. Effects of PACAPs on the increase in intracellular Ca(2+) concentration ([Ca(2+)](i)) in retinal neurons was investigated using the Ca(2+) image analyzing system with fura-2. The cAMP contents and the mitogen-activated protein (MAP) kinase activity in retinal cultures were measured by radioimmunoassay. Concomitant application of PACAPs (10 nM-1 microM) with glutamate (1 mM) for 10 min inhibited the delayed death of retinal neurons, which was observed 24 h after glutamate (1 mM) treatment in a dose-dependent manner. Protection by PACAPs (100 nM) against glutamate-induced neurotoxicity was antagonized by PACAP6-38 (1 microM), a PACAP antagonist, and H-89 (1 microM), a protein kinase A (PKA) inhibitor. However, PACAPs did not affect the glutamate-induced increase in [Ca(2+)](i), but PACAPs (1-100 nM) increased the cAMP levels in a dose-dependent manner. In addition, activation of MAP kinase by PACAP38 (1 microM) was inhibited by simultaneous application with H-89 (1 microM). These findings suggest that PACAPs attenuate glutamate-induced delayed neurotoxicity in cultured retinal neurons by activating MAP kinase through the activation of cAMP-stimulated PKA.

Contraction of Urinary Bladder by Central Norepinephrine Originating in the Locus Coeruleus

Studies were performed to elucidate the role of the locus coeruleus, which is rich in norepinephrine-containing cell bodies, in vesical function using alpha-chloralose anesthetized cats. Stimulation of the locus coeruleus caused contraction of the urinary bladder, which was not affected by transection of the bilateral hypogastric nerves, but blocked by intravenous administration of hexamethonium, a ganglion blocking agent. In animals with transected hypogastric nerves, the locus coeruleus-induced contraction was inhibited by intrathecal administration of phentolamine (alpha-blocker) and prazosin (alpha 1-blocker), but not affected by intrathecal sotalol (beta-blocker) or yohimbine (alpha 2-blocker). In animals treated with reserpine, the locus coeruleus-induced contraction was enhanced by intravenous application of L-dopa, a precursor of norepinephrine. These results suggest that norepinephrine derived from the locus coeruleus activated preganglionic neurons in the sacral intermediolateral nuclei via alpha 1-receptors, thereby producing urinary bladder contraction.

Locus coeruleus-induced inhibition of dorsal cochlear nucleus neurons in comparison with lateral vestibular nucleus neurons.

The effects of conditioning stimulation of the locus coeruleus (LC) on the neuron activity of dorsal cochlear nucleus (DCN), which is rich in noradrenergic nerve terminals, were compared with those on the lateral vestibular nucleus (LVN), devoid of such terminals, to determine whether or not noradrenaline is responsible for the LC-induced inhibition. The conditioning stimuli applied to the LC had no effect on either the field potential or the spike generation of mono- and polysynaptic neurons in the LVN elicited by VIIIth cranial nerve stimulation. In contrast, the spike firing of the DCN neurons with VIIIth cranial nerve stimulation was significantly inhibited by LC conditioning stimulation. The inhibition of spike generation was mainly observed in the DCN neurons which fired spikes with a longer latency. The inhibition of DCN neurons by LC conditioning stimulation did not occur in the cats pretreated with reserpine; however, a rapid recovery of the inhibition was produced by intraventricular application of noradrenaline. These results are in good agreement with the histochemical findings and support our previous conclusion that noradrenaline acts as an inhibitory transmitter or modulator on the nuclei where noradrenergic nerve terminals derived from the LC are located. In addition, the vestibular input in the primary relay nucleus is apparently not regulated by noradrenaline originating in the LC.

Separation of antiepileptogenic and antiseizure effects of levetiracetam in the spontaneously epileptic rat (SER)

Summary:  Purpose: The long‐lasting antiseizure effects of levetiracetam (LEV) have been observed in the spontaneously epileptic rat (SER) that expresses both tonic and absence‐like seizures. Furthermore, the antiepileptogenic effects of LEV in addition to antiseizure effects have been reported in the amygdala‐kindling model in rats. This suggests that the long‐lasting seizure protection of LEV may be at least partly due to its antiepileptogenic effects. Therefore this study aimed to differentiate the antiseizure and potential antiepileptogenic effects of LEV by administering LEV continuously to SERs before the appearance of any seizure expression.

Mediation of micturition reflex by central norepinephrine from the locus coeruleus in the cat

We examined whether norepinephrine originating in the locus coeruleus mediates the micturition reflex in anesthetized cats. 6-Hydroxydopamine, a catecholamine neurotoxin, injected bilaterally into the locus coeruleus markedly decreased catecholamine fluorescence in the lesioned area and induced urinary retention after 72 to 84 hr. At this time, there was no or only slight contraction of the urinary bladder induced by its distension, while the contraction was noted before the treatment. However, phenylephrine, an alpha 1-receptor agonist, applied intrathecally in 6-hydroxydopamine-treated animals induced moderate bladder contraction. In sham-operated animals, the bladder contraction on its distension was inhibited by intrathecally applied prazosin, an alpha 1-receptor antagonist. Thus, in the micturition reflex, norepinephrine derived from the locus coeruleus acts on the alpha 1-adrenergic receptors in the sacral cord, and induces urinary bladder contraction via activation of the sacral parasympathetic preganglionic neurons.