Morphology and electrophysiology of ventral mesencephalon nerve cell cultures.

In primary neuron cultures obtained from ventral mesencephalon of mouse fetuses, approximately 10-30% of the neurons were dopaminergic, as demonstrated by a rapid glyoxylic acid histofluorescence procedure, and another 10-30% were GABAergic as demonstrated by autoradiography. Resting membrane potentials averaged -58 mV and input resistances averaged 188 M omega. Action potential (AP) firing patterns were of 3 types: in 49% of cells, depolarizing current elicited bursts of APs of constant amplitude, duration, and interspike interval (Type 1); in 44% of cells, bursts consisted of APs of decreasing amplitude, increasing duration, and increasing interspike interval (Type 2); and in 7% of cells, bursts were initiated by a single high amplitude, short duration AP followed by a series of lower amplitude longer duration APs that progressively increased in amplitude and decreased in duration and interspike interval (Type 3). Calcium APs of two distinct types, differing in duration and rate of rise, were observed when cultures were exposed to tetrodotoxin. Abundant postsynaptic activity was recorded. Simultaneous intracellular recording between pairs of cells demonstrated reciprocal innervation. The neurotransmitter antagonists haloperidol, bicuculline, naloxone, atropine, hexamethonium and pirenzepine affected synaptic activity and/or resting membrane potential of some of the cultured neurons.

Acetylcholine responses in synaptically active neurons in mouse ventral mesencephalon cultures.

Acetylcholine (ACh) sensitivity was examined in neurons in dissociated cultures from fetal ventral mesencephalon (14-16 days gestational age); many neurons probably originate in substantia nigra. Direct responses to iontophoretically applied ACh were recorded by intracellular microelectrodes in 53 cells and indirect responses were recorded from 58 synaptically connected cells. Cultures (24-50 days old) were studied in growth media enriched with serum (10%) and calcium (6.8 mM). Direct responses: ACh caused a slow depolarization (seconds) in 94% (50/53); in 30% of these cells (15/50) the depolarization exceeded action potential threshold. Hyperpolarizing responses occurred in 4% (2/53) and another cell showed reduced action potential firing. Indirect responses: Application of ACh to adjacent cells caused an increase in postsynaptic potential (PSP) activity which was excitatory in 66% (38/58) and inhibitory in 17% (10/58). A reduced level of PSP activity occurred in 7% (4/58) of cells exhibiting excitatory PSPs and in 10% (6/58) of cells with inhibitory connections. Indirect responses were blocked by tetrodotoxin. ACh receptor types: Responses to ACh were predominantly muscarinic (77%, 10/13). Nicotinic responses were present in the remaining 23% (3/13).

Human cytomegalovirus infection of human placental explants in culture: histologic and immunohistochemical studies.

The induction of human cytomegalovirus infection in human first-trimester placentas was studied with a placental explant culture model. Replication and/or release of human cytomegalovirus in placental explant cultures did not occur at any time from 1 to 10 days after infection when examined by plaque assay and analyses of extracted deoxyribonucleic acids. In contrast, typical human cytomegalovirus-induced histopathologic lesions bearing human cytomegalovirus antigens were consistently localized in the trophoblastic cells covering placental villi. These data clearly demonstrate that placental cells are permissive of latent and/or abortive human cytomegalovirus infection in vitro. Our results support the hypothesis that during human cytomegalovirus infection of pregnant women, maternal viremia or intrauterine infection results in latent human cytomegalovirus infection of placental cells that may persist during the course of pregnancy.

Interactions between serotonin and cisapride on myenteric neurons.

Intracellular recording methods were used to investigate the interactions between serotonin (5-HT) and cisapride on myenteric neurons of guinea-pig small intestine. Serotonin had three actions on the neurons. One was a slowly rising depolarization associated with increased input resistance and discharge of spikes that lasted six or more times longer than the duration of the 5-HT application. The second action was a transient depolarization associated with decreased input resistance and brief discharge of spikes. This response desensitized quickly and could be evoked only at intervals of 2 to 3 min. The third action of 5-HT was presynaptic inhibition of acetylcholine release at nicotinic synapses. Cisapride reduced or abolished both the prolonged and transient responses to 5-HT. The threshold concentration for reduction of the responses was 0.1 microM and the responses were abolished at 1.0 to 10 microM. Cisapride suppressed stimulus-evoked slow excitatory postsynaptic potentials (EPSPs) in the same cells for which cisapride blocked the prolonged responses to 5-HT. There were no effects of cisapride on resting electrical behavior or spike generation. Cisapride reduced the amplitude of fast cholinergic EPSPs, suggesting that it behaved as an agonist at the presynaptic serotonergic receptors.

Intracellular study of effects of histamine on electrical behaviour of myenteric neurones in guinea-pig small intestine.

The actions of histamine on myenteric neurones were investigated with intracellular recording methods in guinea-pig small intestine. The actions of histamine at the ganglion cell soma were: membrane depolarization, increased input resistance, suppression of post-spike hyperpolarizing potentials, augmented excitability and repetitive spike discharge. Excitability was enhanced also at spike initiation sites remote from the cell body. Both H1, and H2, receptors were involved in the response to histamine. Dimaprit mimicked the responses to histamine in 80% and 2-methylhistamine in 50% of the trials. Cimetidine was an antagonist for histamine in 82% and for dimaprit in all of the trials. Pyrilamine blocked the actions of histamine in 59% of the cells and always blocked the action of 2-methylhistamine. Histamine mimicked slow synaptic excitation in the neurones, but was ruled out as a neurotransmitter for the slow excitatory post-synaptic potential (e.p.s.p.). Histamine either did not affect the responses to 5-hydroxytryptamine, substance P and acetylcholine or it potentiated the responses to these putative neurotransmitters for slow synaptic excitation. The results support the possibility that histamine released from mast cells by circulating peptidergic messengers, by neurotransmitters or during anaphylaxis could influence enteric nervous function.

Neuronal generation of the leech swimming movement.

The swimming movement of the leech is produced by an ensemble of bilaterally symmetric, rhythmically active pairs of motor neurons present in each segmental ganglion of the ventral nerve cord. These motor neurons innervate the longitudinal muscles in dorsal or ventral sectors of the segmental body wall. Their duty cycles are phase-locked in a manner such that the dorsal and ventral body wall sectors of any given segment undergo an antiphasic contractile rhythm and that the contractile rhythms of different segments form a rostrocaudal phase progression. This activity rhythm is imposed on the motor neurons by a central swim oscillator, of which four bilaterally symmetric pairs of interneurons present in each segmental ganglion appear to constitute the major component. These interneurons are linked intra- and intersegmentally via inhibitory connections to form a segmentally iterated and inter-segmentally concatenated cyclic neuronal network. The network appears to owe its oscillatory activity pattern to the mechanism of recurrent cyclic inhibition.

The distribution of acetylcholine receptors on muscle fibres of regenerating salamander limbs.

1. Muscle fibres developing during limb regeneration were examined for responsiveness to acetylcholine (ACh) applied iontophoretically along the membrane. 2. Fibres which were uninnervated as well as those with non-transmitting synapses had high over-all sensitivities, with only minor variations along their length. 3. Functionally innervated fibres in which depolarization did not yet elicit action potentials had high over-all sensitivities, even when the synaptic potentials ahd amplitudes of 40-50 mV. In these the membrane in the vicinity of synapses tended to have sensitivities above the background level. 4. Upon the appearance of action potentials, several weeks after fibre innervation, the responsiveness to ACh began to decline in synapse free regions of the membrane. In mature muscle the sensitivity to ACh is restricted to sites of synaptic contact.