Action of morphine on rat cortical neurons intracellularly recorded in vivo: evidence for an excitatory postsynaptic effect which is naloxone insensitive

The action of morphine, applied either iontophoretically (40-200 nA balanced current) or systemically (5-10 mg/kg, intraperitoneally) to rat cortical neurons, was investigated in vivo, using intracellular electrodes. Morphine increased the apparent input resistance and increased the number of both spontaneous and evoked action potentials. Several cells, which normally generated single spikes, generated bursting potentials; neurons with bursting activity increased their activity. Naloxone, iontophoretically or systemically applied, did not reverse or prevent the morphine-induced excitation. The iontophoretic administration of cadmium suggested that the effects of morphine were due, at least in part, to a postsynaptic site of action. It is suggested that the increase of cellular excitability induced by morphine could contribute to its production of seizures in cortex.     

吗啡对离体兔小肠平滑肌电变化及环肌和纵肌收缩活动的影响

本实验同时观察吗啡对离体兔十二指肠平滑肌电活动,肠腔内压变化以及纵肌收缩活动三方面的影响。结果显示:吗啡可使离体兔十二指肠节律性环肌收缩加强,纵肌收缩减弱,以及平滑肌峰电位幅度,数量和峰电位发生率增加。吗啡的小肠兴奋作用可被纳洛酮或阿托品阻断。吗啡对纵肌的抑制作用不被心得安阻断,但可被酚妥拉明减弱。实验结果表明吗啡对兔小肠环肌和纵肌作用相反,它兴奋环肌,抑制纵肌。吗啡是通过肠道阿片受体起作用的。乙酰胆碱参与了吗啡对小肠的作用。     

Behavioral and electrophysiological evidence for tolerance to continuous morphine administration into the ventrolateral periaqueductal gray

Repeated microinjections of morphine into the ventrolateral periaqueductal gray (vPAG) produce tolerance to the antinociceptive effect of morphine [Behav Neurosci 113 (1999) 833]. These results may be a direct effect of morphine on cells within the vPAG or be caused by cues linked to the microinjection procedure (i.e. associative tolerance). The objective of this paper was to determine whether continuous administration of morphine into the vPAG (i.e. no cues) would produce tolerance. Tolerance was assessed by measuring changes in behavior and changes in the activity of neurons in the rostral ventromedial medulla (RVM), the primary output target of the PAG. Rats were implanted with an osmotic minipump that released morphine (2.5 or 5 microg/h) or saline into the vPAG continuously. Continuous administration of morphine produced an increase in hotplate latency when measured 6 h after initiation of treatment. Tolerance to this antinociception was evident within 24 h. After 3 days, rats were anesthetized and the activity of RVM neurons was assessed. Although acute morphine administration into the RVM inhibits the activity of RVM on-cells and enhances the activity of off-cells, these neurons appeared normal following 3 days of continuous morphine administration. Systemic naloxone administration produced hyperalgesia that was associated with a marked increase in on-cell activity and a complete cessation of off-cell activity. The loss of morphine inhibition of nociception, measured behaviorally and electrophysiologically, demonstrates that tolerance is caused by a direct action of morphine on vPAG neurons.     

Opioid peptides as possible neuromodulators of the afferent synaptic transmission in the frog semicircular canal.

Vestibular receptors of the frog, Rana temporaria, were examined for the effect of bath-applied opioid peptide leu-enkephalin, its synthetic analogue dalargin and the specific opiate antagonist naloxone. Multiunit afferent activity of the whole vestibular nerve was recorded in an in vitro preparation. Leu-enkephalin (0.005-100 nM) and dalargin (0.1-100 nM) depress the resting discharge frequency. Naloxone (10 nM-1 microM) antagonizes responses induced by leu-enkephalin and dalargin that suggests a specific action of opioid peptides. Leu-enkephalin and delargin inhibit the excitatory action of L-glutamate. The effects of opioid peptides on L-glutamate-induced responses are unaffected by Co2+ block of transmitter release from hair cells that could speak in favour of the postsynaptic nature of these responses. At the same time, the other possible site of action of opioid peptides, such as efferent system, can not be excluded. The results indicate that opiate receptors are present in hair cells and that the neurotransmitter L-glutamate is involved in opiate action at the peripheral vestibular system of the frog. We suggest that opioid peptides may act as a neuromodulator in this system.     

The actions of histamine in cat and rabbit superior cervical ganglia

Depolarization of the cat superior cervical ganglion (SCG) evoked by histamine was antagonized by mepyramine. Histamine-induced depolarization, indicated by changes in the negative and positive afterpotentials of ganglionic action potentials, was decreased by pretreatment with aminophylline. In the cat SCG, histamine evoked stimulus bound decremental oscillatory potentials (SBDOP). In the rabbit SCG, only depolarization and no SBDOP were observed after the administration of histamine. Histamine-induced SBDOP were increased by isoprenaline and decreased by Leu-enkephalin pretreatment. Our results indicate that histamine evokes depolarization and an increase in excitability of the cat SCG which seems to be mediated by H₁-receptors.     

Electrical behaviour in a line of anterior pituitary cells (GH cells) and the influence of the hypothalamic peptide, thyrotrophin releasing factor

Anterior pituitary cells of the GH line, which secrete prolactin spontaneously, showed spontaneous action potential activity. Thyrotrophin releasing factor, which increases secretion in these cells, caused a prompt increase of action potential frequency. Potassium, another secretagogue, depolarized the cells and sometimes initiated a burst of action potentials at the onset of this effect. The action potentials persisted in tetrodotoxin-containing and Na-free media, but were suppressed by the Ca-channel blocker, methoxyverapamil. Moreover, elevating the extracellular Ca²⁺ concentration increased the amplitude of the action potentials. These action potentials therefore have a prominent Ca component. This endows them with a particular interest since secretory activity of these cells is known to be dependent on extracellular Ca²⁺. Ba²⁺, which can substitute for Ca²⁺ in maintaining secretion, also substituted for Ca²⁺ in the maintenance of the action potentials. In addition, Ba²⁺ prolonged action potentials remarkably: tetraethylammonium was less effective in this regard.The several parallels between known secretory behaviour and electrical phenomena encourage the view that analysis of electrical activity in anterior pituitary cells may provide useful clues to events involved in stimulus-secretion coupling and in the secretory control exerted by the brain.     

Morphine induces a spontaneous and evoked bursting activity in rat cortical neurons by adding a postsynaptic active mechanism to the synaptic input: an intracellular study in vivo

The action of morphine on spontaneous and stimulus-evoked postsynaptic potentials was investigated in rat cortical neurons recorded intracellularly in vivo. Iontophoretically applied, morphine increased supra-threshold evoked depolarizing postsynaptic potentials inducing bursts of spikes, but only slightly increased weak (subthreshold) potentials. Spontaneous excitatory postsynaptic potentials were affected in a similar way, but their frequency did not change. Inhibitory postsynaptic potentials were only subsequently modified. Membrane hyperpolarization, induced by negative current injection, counteracts the morphine-induced burst generation. We suggest that the action of this alkaloid on threshold postsynaptic events involves a voltage-dependent mechanism, which may be triggered by synaptic currents.     

Growth-inhibiting extracellular matrix proteins also inhibit electrical activity by reducing calcium and increasing potassium conductances

Inhibitionof neurite sprouting and electrical activity by extracellular matrix (ECM) glycoproteins was studied during neurite regeneration by using anterior pagoda (AP) neurons of the leech. Adult isolated neurons were plated in culture inside ganglion capsules, which among many ECM proteins, contain a group of inhibitory peanut lectin- (PNA) binding glycoproteins. These proteins inhibit neurite production and contribute to the formation of a bipolar outgrowth pattern by AP neurons. Addition of PNA lectin to the culture medium to block the inhibitory effects of ECM glycoproteins induced an increase of neurite sprouting, the loss of the bipolar pattern, and also an increase in the amplitude and duration of action potentials evoked by intracellular current injection. PNA lectin had independent effects on neurite sprouting and electrical activity, since there was no correlation between the total neurite length and the amplitude of the action potentials. Moreover, action potentials were increased by the presence of PNA lectin even in neurons that did not grow. The changes induced by PNA lectin on the active conductances underlying the action potentials were estimated by quantitative model simulations. We predict that the increases in the amplitude and duration of the action potential induced by PNA lectin were due to an increase in a calcium conductance and a reduction in the delayed rectifier potassium conductance. Our results suggest that inhibitory ECM glycoproteins may use independent signaling pathways to inhibit neurite sprouting and electrical activity. These proteins affect the action potential by changing the proportion of inward and outward active conductances.     

Enigmatic action of ciclosporine A on the naloxone-precipitated morphine withdrawal syndrome in mice

Various alterations of the immune system have recently been reported to attenuate the severity of morphine withdrawal. The effect of the immunosuppressive agent ciclosporine A on the naloxone-induced morphine withdrawal syndrome in the chronically dependent mouse was investigated. Ciclosporine A significantly suppressed stereotyped behaviour such as jumping and forepaw treading while wet shakes were potentiated. Withdrawal diarrhoea was diminished as a consequence of a promotive action of ciclosporine A on the intestine. A ciclosporine derivative, which is devoid of immunosuppressive activity, had no influence on withdrawal signs. The attenuating effect of ciclosporine A was observed at a dose of 20 mg/kg i.p., which is not regarded as immunosuppressive in the mouse. It was also effective in animals lacking an intact immune system as a result of a genetic T-cell defect (nude mouse) or after selective ablation by whole body irradiation. Nude mice and irradiated normal mice developed dependence on morphine to the same extent as normal animals, as could be derived from the severity of their withdrawal signs. These results suggest that an intact immune system is not a necessary prerequisite for ciclosporine A to attenuate morphine withdrawal and that its action may be attributable to mechanisms other than immunosuppression. It is possibly a result of a direct effect of ciclosporine A on the central nervous system structures involved in the behavioural expression of the opiate withdrawal syndrome.     

Alterations in opioid system of the rat brain after cat odor exposure

The effect of cat odor exposure was studied on morphine-induced increase of exploratory behavior and on the expression of opioid genes in forebrain structures of male Wistar rats. Treatment with morphine (1 mg/kg) induced a significant increase in exploratory behavior in an unfamiliar environment in rats. Previous exposure of animals to cat odor completely abolished this stimulating action of mu-opioid receptor agonist on exploratory activity. Cat odor exposure induced a significant increase in the expression of pro-opio-melanocortin (POMC) and mu-opioid receptor (MOR) genes in the brain structures related to anxiety and motivation. This study clearly demonstrates that cat odor exposure increases the activity of opioid system in rat forebrain structures.     

Comparison of the hyperglycaemic effect of adrenaline and morphine introduced into the liquor space.

1. In unanaesthetized cats a comparison is made of the hyperglycaemic effects of adrenaline and morphine, when injected or infused through chronically implanted cannulae, into different regions of the cerebral ventricles or of the subarachnoid space, in order to determine their sites of action. 2. On injection into the cerebral ventricles both adrenaline and morphine have to reach the subarachnoid space beneath the ventral surface of the brain stem before they can exert their hyperglycaemic effect. The adrenaline has to reach the region rostral to the pons, i.e. the fossa interpeduncularis, and the morphine the region caudal to the trapezoid bodies. These conclusions are based on the following findings. 3. When adrenaline (55 mug) and morphine (0-75mg) were infused into one or other of these two regions, adrenaline produced strong hyperglycaemia on infusion into the fossa interpeduncularis, but had scarcely any hyperglycaemic effect on infusion into the region caudal to the trapezoid bodies. The reverse result was obtained with morphine. 4. It is concluded that the adrenaline hyperglycaemia is mainly a peripheral effect. It occurs after the adrenaline has been absorbed into the blood stream from the fossa interpeduncularis but an additional central component, an action on brain stem structures reached from the fossa interpeduncularis, cannot be excluded. The morphine hyperglycaemia is a central effect due to an action on superficial structures of the ventral surface of the medulla oblongata, caudal to the trapezoid bodies.     

Transmission from perivascular inhibitory nerves to the smooth muscle of the guinea-pig taenia coli

1. Membrane potential changes of the smooth muscle cells of the taenia coli were recorded during stimulation of the perivascular inhibitory nerves.2. Some spontaneous action potentials were preceded by a slow pacemaker-like potential. Others began at or near the maximum level of the membrane potential and were not preceded by pacemaker-like potentials.3. There were no changes in the membrane potential of smooth muscle cells when the inhibitory nerves were stimulated with a single pulse. Stimulation at frequencies greater than 5 pulses/sec caused a hyperpolarization of the smooth muscle membrane. This resulted in a decrease in spike frequency and relaxation.4. When the frequency of stimulation of the inhibitory nerves was increased there was an increase in the amplitude and rate of rise of the hyperpolarization and a decrease of the latency. The latency varied from 150 to 300 msec, and the largest hyperpolarization recorded was 16 mV.5. The effect of the hyperpolarization due to nerve stimulation in cells showing pacemaker-like activity was to increase the level of the membrane potential at which the action potentials began and to increase the membrane potential to which the action potentials repolarized. Action potentials which occurred during hyperpolarizations of the membrane had greater rates of rise and fall and larger amplitudes than did the action potentials which occurred before hyperpolarization.6. The effect of the hyperpolarization due to nerve stimulation in cells which did not show pacemaker-like activity depended on the amplitude of the hyperpolarization. Small hyperpolarizations exposed small depolarizations of the membrane which occurred when an action potential would normally have been expected. Large hyperpolarizations blocked the action potentials entirely.7. Action potentials did not begin firing again at the normal rate immediately after stimulation ceased. The time taken for the rate of firing of action potentials to return to normal increased with increasing frequency of stimulation.8. The hyperpolarization in response to perivascular inhibitory nerve stimulation was blocked by guanethidine and bretylium.     

A microiontophoretic study of morphine on single neurons in the rat globus pallidus

The microiontophoretic application of morphine to single globus pallidus neurons in morphine naive rats resulted in a depression of both spontaneous and glutamate-evoked firing of these neurons. Seventy percent of all pallidal neurons on which morphine was tested exhibited some degree of depression of neuronal discharge; no excitatory effect was observed to morphine on any pallidal neurons. The depressant effects of morphine, but not those of dopamine, could be antagonized by the microiontophoretic application of the morphine antagonist naloxone, indicating that the morphine-induced depression of pallidal neuronal activity was a specific morphine action. This effect of morphine is most likely mediated by the opiate receptors on pallidal neurons that have been proposed to function in enkephalinergic-mediated synaptic transmission.     

Apparent dissociation between saccadic eye movements and the firing patterns of premotor neurons and motoneurons

Saccadic eye movements result from high-frequency bursts of activity in ocular motoneurons. This phasic activity originates in premotor burst neurons. When the head is restrained, the number of action potentials in the bursts of burst neurons and motoneurons increases linearly with eye movement amplitude. However, when the head is unrestrained, the number of action potentials now increase as a function of the change in the direction of the line of sight during eye movements of relatively similar amplitudes. These data suggest an apparent uncoupling of premotor neuron and motoneuron activity from the resultant eye movement.     

Quinine suppresses extracellular potassium transients and ictal epileptiform activity without decreasing neuronal excitability in vitro

The effect of quinine on pyramidal cell intrinsic properties, extracellular potassium transients, and epileptiform activity was studied in vitro using the rat hippocampal slice preparation. Quinine enhanced excitatory post-synaptic potentials and decreased fast- and slow-inhibitory post-synaptic potentials. Quinine reduced the peak potassium rise following tetanic stimulation but did not affect the potassium clearance rate. Epileptiform activity induced by either low-Ca(2+) or high-K(+) artificial cerebrospinal fluid (ACSF) was suppressed by quinine. The frequency of spontaneous inter-ictal bursting induced by picrotoxin, high-K(+), or 4-aminopyridine was significantly increased. In normal ACSF, quinine did not affect CA1 pyramidal cell resting membrane potential, input resistance, threshold for action potentials triggered by intracellular or extracellular stimulation, or the orthodromic and antidromic evoked population spike amplitude. The main effects of quinine on intrinsic cell properties were to increase action potential duration and to reduce firing frequency during sustained membrane depolarizations, but not at normal resting membrane potentials. This attenuation was enhanced at increasingly depolarized membrane potentials.These results suggest that quinine suppresses extracellular potassium transients and ictal activity and modulates inter-ictal activity by limiting the firing rate of cells in a voltage-dependent manner. Because quinine does not affect 'normal' neuronal function, it may merit consideration as an anticonvulsant.     

REM sleep deprivation antagonizes morphine-induced akinesia and catalepsy

An examination was made of the effect of REM sleep deprivation (REMSD) on some forms of altered motor activity, such as akinesia and catalepsy, induced by intraperitoneal (i.p.) or intracerebroventricular (i.c.v.) administration of morphine in adult, male Wistar rats. Administration of morphine (25 mg/kg i.p.) induced an akinetic-cataleptic syndrome and decreased spontaneous vertical motor activity (SVMA) in animals allowed undisturbed sleep. REMSD decreased the morphine-induced akinesia and catalepsy that are known to be mediated by an inhibitory mu-opiate system. The locomotor depressant action of morphine was converted to excitation (manifested as increased SVMA and hopping behavior) by REMSD. Similarly, decreased motor activity following i.c.v. administration of morphine (25 micrograms) was replaced by excitation in the form of jumping behavior after REMSD. Naltrexone (1 mg/kg i.p.) blocked the akinetic and cataleptic effects, but not the excitatory effects, of morphine. It is suggested that REMSD is associated with a functional insufficiency of an inhibitory mu-opiate system, thus unmasking the excitatory morphine effects. The proposed insufficiency of an endogenous opioid system might explain an increase in neuronal excitation during REMSD and the therapeutic effect of REM deficiency in some types of depression.     

Naloxone-induced depolarization and synaptic activation of myenteric neurons in morphine-dependent guinea pig ileum

To investigate the cellular basis of opiate dependence, intracellular microelectrodes were used to record from both electrophysiologically defined classes of neurons (S and AH) in myenteric plexus longitudinal muscle preparations from morphine pretreated guinea pigs. These preparations responded to naloxone with the characteristic contraction of the longitudinal smooth muscle, indicative of morphine dependence. Depolarization in response to naloxone was observed in 42% of S neurons, but there were no consistent changes in input resistance. In some cells the depolarization was reduced or abolished after blockade of synaptic transmission, suggesting that it was due in part to the release of an excitatory transmitter producing a slow depolarization in the impaled neuron. Synaptic activation of S neurons during withdrawal was further indicated by the observation that fast postsynaptic potentials appeared after abrupt displacement of morphine from its receptors by naloxone. Morphine withdrawal, therefore, involves both the final motor neurons and interneurons. During naloxone-induced withdrawal, 25% of S neurons discharged action potentials. In contrast, no action potentials were discharged in AH neurons. Furthermore, naloxone did not alter the resting membrane potential, input resistance, soma action potential configuration, or slow hyperpolarization following a soma spike in AH neurons. The specificity of the withdrawal response for S neurons and the relatively small proportion of neurons involved suggests that morphine withdrawal occurs in quite specific neuronal circuits in the myenteric plexus.     

[3H]Leu-enkephalin binding following chronic swim-stress in mice

Warm water swimming produces in mice an opiate-like antinociceptive response. Chronic swimming produces tolerance to the antinociceptive response and, depending on the schedule, cross-tolerance with morphine and naloxone intensified withdrawal signs. Low affinity [³H]Leu-enkephalin binding to brain homogenates at low temperature was significantly reduced in acutely swum mice and chronically swum mice whether or not they were swum. Preincubation at 37°C abolished all between-group differences. Results following chronic swimming were similar whether or not the schedule produced morphine cross-tolerance. These results were discussed in terms of the interpretation that reduced binding reflects increased in vivo occupation of opioid binding sites.     

Augmenting action of nicotine on population spikes in the dentate gyrus of the guinea pig

Effects of nicotinic cholinergic agents on field potentials recorded from the dentate gyrus were studied in thin transverse sections of the hippocampus of the guinea pig. Nicotine augmented the population spike elicited by the second stimulus of a paired stimulation to the molecular layer. The threshold concentration of nicotine to cause this effect was 5–10 μM. The augmentation of the spike was not accompanied by an increase in the rising slope of population excitatory postsynaptic potentials, and was not observed in the presence of bicuculline. Carbamylcholine had a weak and inconsistent effect. D-tubocurarine and mecamylamine also augmented the population spike. The action of nicotine was blocked by hexamethonium. These results suggest that nicotine facilitates the generation of action potentials in granule cells by depressing inhibitory processes, and that properties of nicotinic cholinergic receptors are different in different subfields of the hippocampal formation, presumably reflecting the diversity of the receptors.     

Valproate attenuates the development of morphine antinociceptive tolerance

Morphine is a potent opioid analgesic. Repeated administration of morphine induces tolerance, thus reducing the effectiveness of analgesic treatment. Although some adjuvant analgesics can increase morphine analgesia, the precise molecular mechanism behind their effects remains unclear. Opioids bind to the mu opioid receptor (MOR). Morphine tolerance may be derived from alterations in the intracellular signal transduction after MOR activation. Chronic morphine treatment activates glycogen synthase kinase 3β (GSK3β), whose inhibition diminishes morphine tolerance. Valproate is widely prescribed as an anticonvulsant and a mood stabilizer for bipolar disorders because it increases the amount of γ-aminobutyric acid (GABA) in the central nervous system. Although the activation of GABAergic neurons may be responsible for the chief pharmacologic effect of valproate, recent studies have shown that valproate also suppresses GSK3β activity. We examined the effect of valproate on the development of morphine antinociceptive tolerance in a mouse model of thermal injury. Mice were treated with morphine alone or with morphine and valproate twice daily for 5 days. The resulting antinociceptive effects were assessed using a hot plate test. While mice treated with morphine developed tolerance, co-administration of valproate attenuated the development of tolerance and impaired the activation of GSK3β in mice brains. Valproate alone did not show analgesic effects; nevertheless, it functioned as an adjuvant analgesic to prevent the development of morphine tolerance. These results suggest that the modulation of GSK3β activity by valproate may be useful and may play a role in the prevention of morphine tolerance.