Robert B. Hill

THE EFFECTS OF CERTAIN NEUROHUMORS AND OF OTHER DRUGS ON THE VENTRICLE AND RADULA PROTRACTOR OF BUSYCON CANALICULATUM AND ON THE VENTRICLE OF STROMBUS GIGAS

1. The hearts of Busycon canaliculatum and Strombus gigas were found to respond to applied neurohumors as do the myogenic hearts of other gastropods. Acetyicholine was cardio-inhibitory, and 5-hydroxytryptamine was cardio-acceleratory, in concentrations low enough to suggest that they might be the normal regulatory neurohumors.2. The Busycon canaliculatum radula protractor was contracted by acetylcholine, and could then be relaxed rhythmically by 5-hydroxytryptamine, tryptamine, and adrenalin, all of which raise the tonus of the ventricle.

Ionic dependence of the response to acetylcholine of a molluscan buccal muscle: The radular protractor of Busycon canaliculatum

Abstract 1. We studied the ionic dependence of the response of a molluscan phasic smooth muscle to applied acetycholine (ACh). 2. In sodium-free medium, ACh depolarization was eventually greatly reduced and contracture-force was nearly abolished. 3. Potassium-free medium did not affect ACh depolarization or contracture-force. 4. Exposure to low-calcium medium did not lead to a reduction in amplitude of ACh depolarization but force in low-calcium medium was eventually reduced to zero. 5. These results are consistent with involvement of inward Na+ current in depolarization by ACh and of Ca2+ in excitation-contraction coupling.

Use of a sucrose gap apparatus to record electrical responses of gastropod radular protractor muscle to fmrfamide

Abstract 1. 1. Auxotonic contractions of the radular protractor muscle (RPM) of Busycon contrarium have often been used for bioassay of FMRFamide (Fa). Until now, however, sucrose gap recording of electrical responses of the RPM of Busycon canaliculatum has invariably been accompanied by isometric recording of contractions. 2. 2. We find that isometric responses of the RPM of B. canaliculatum seem to be less sensitive detectors of Fa than the auxotonic responses of the RPM of B. contrarium , which are used for bioassay. Therefore, we have now combined sucrose gap recording of electrical responses with auxotonic recording of contractions. 3. 3. The most interesting outcome of this study was the discovery that the responses of RPMs to FMRFamide are accompanied by very little depolarization, compared to their responses to potassium (K) or acetylcholine (ACh). 4. 4. We have also found that low concentrations of Fa induce very weak, slow contractions for which auxotonic recording provided more reliable detection of threshold responses than did isometric recording. However, strength of contraction (but not depolarization) increased remarkably with concentration.

ANATOMY OF THE INNERVATION AND NEUROMUSCULAR JUNCTIONS OF THE RADULAR PROTRACTOR MUSCLE OF THE WHELK, BUSYCON CANALICULATUM (L.)

1. A detailed and integrated picture of the whole buccal innervation of Busycon canaliculatum is presented.2. The results of our observations on the fine structure of the nerves and of the neuromuscular junctions in the radular protractor are reported and discussed. Each radular protractor muscle is innervated by a nerve arising from the cerebrobuccal connective. The nerve trunk enters the proximal end of the muscle and runs parallel to the long axis of the muscle bundle. A layer of connective tissue and epineural muscle cells surrounds the trunk. Subdivisions of the main nerve trunk branch laterally into the muscle bundle losing their epineural muscle coat. Further subdivisions of the branches produce single axons which can be observed among the muscle cells, but no specialized motor nerve endings were observed. Within the nerve endings are two types of synaptic vesicles: agranular (clear) and granular (dense). The granular vesicles are larger, ranging in diameter from 800 to 1350 A (mean 970 A). The cle...

Unusual responses of proboscis muscles ofBusycon canaliculatum to some calcium antagonist agents

SummaryK- and ACh-induced responses of the radular sac, odontophore retractor, and radular retractor muscles ofBusycon canaliculatum were found to be strongly dependent upon [Ca]0. Diltiazem had strong positive inotropic and chronotropic actions on fast twitch activity in the odontophore retractor and radular protractor muscles. K-induced tonic force in these muscles was partly inhibited by diltiazem but only at very high concentrations. ACh responses in all muscles were eliminated by diltiazem. Nifedipine enhanced fast twitches and tonic force in response to high K, and induced persistent spontaneous fast twitch discharges. Nifedipine inhibited ACh-induced tonic force, but induced rhythmic bursts of fast twitches persisting long after nifedipine washout. Verapamil strongly inhibited K- and ACh-induced tonic force in all three muscles at high concentration, but stimulated fast twitch responses and converted ACh contractures into fast twitch activity. Sucrose gap studies showed that nifedipine and diltiazem reduced K- and ACh-induced tension and depolarization. Paradoxically, verapamil reduced K- and ACh-induced tension but significantly enhanced their induced depolarizations. Diltiazem, nifedipine and verapamil did not act like slow Ca channel antagonists in these muscles. This may reflect differences in channel structure between molluscs and mammals, or differences in the cellular calcium release pathways operated by such channels in molluscan and mammalian muscle. These ‘Ca-ant-agonists’ appeared to act as agonists of fast twitch activity in these muscles and antagonists of the ACh-induced calcium release pathway for tonic force development.

Diversity of mechanical responses and their possible underlying mechanisms in the proboscis muscles ofBusycon canaliculatum

SummaryMechanical responses of the radular protractor and retractor, the odontophore retractor and the radular sac muscles ofBusycon canaliculatum were compared. The radular protractor responded to both ACh and high K salines with similar slow, smooth contractures showing no evidence of fast twitch activity. The radular sac, odontophore retractor, and radular retractor muscles responded to low K salines with bursts of fast twitches at a mechanical threshold below that for responses in the radular protractor. With high K salines these three muscles showed inactivation of fast twitch activity and replacement by slow maintained tonic force. With rare exceptions, the ACh responses of all four muscles consisted of slow, maintained tonic contractures with no fast twitch activity, although individual muscles differed in their ACh sensitivity. A scheme is presented to explain the mechanical modus operandi of this complex organ by the co-operative actions of these four physiologically diverse muscles. It is proposed that fast twitch responses depend upon the activity of fast transient Ca channels showing strong voltage sensitivity and ready voltage inactivation. It is proposed that maintained tonic contractures in all the muscles depends upon the activity of slow long-lasting voltage-dependent Ca channels which only open with substantial membrane depolarization. It is suggested that K-induced and ACh-induced responses may activate a similar cellular Ca pool but by different membrane transduction routes.

Modulatory mechanisms in the isolated internally perfused ventricle of the whelk Busycon canaliculatum

1. Isolated cannulated ventricles commenced spontaneous beating on application of perfusion pressure of 10 cm water. Complete hearts showed a fast patterned cyclical rhythm, whereas ventricles devoid of atrial material showed a continuous slow rhythm. 2. Perfused ventricles were inhibited by ACh with a threshold at 10(-8) mol l-1 and arrested at 10(-7) mol l-1, and ventricles under stimulation by 5HT could be arrested by ACh at this concentration. 3. Perfused ventricles were stimulated by 5HT, with threshold at 10(-9) mol l-1 and maximum at 10(-5) mol l-1. Metoclopramide was without affect on 5HT responses, but metitipine and methysergide did inhibit such responses suggesting that the 5HT receptor present possessed mixed properties of the vertebrate 5-HT1 and 5-HT2 receptor subtypes. 4. Ventricles were very sensitive to the excitatory actions of FMRFamide in the 10(-9) to 10(-5) mol l-1 range. Preparations were insensitive to GAPFLRFamide, but SCP-B was modestly excitatory (threshold 10(-7) mol l-1). 5. Preparations were not significantly affected by adenosine, ATP, and guanosine, but GTP was strongly excitatory at 10(-7) mol l-1. 6. 5HT and FMRFamide responses were additive. Preparations responded strongly to the adenylate cyclase activator forskolin and dibutyryl cAMP enhanced spontaneous contractions and 5HT responses, suggesting that the 5HT receptor may operate via a cAMP secondary mechanism. 7. The IP3 inhibitor lithium (10 mmol l-1), caused slight inhibition of FMRFamide responses, suggesting that the receptor to this peptide may operate via IP3 as a second messenger. 8. Neuromodulation in this preparation would appear to involve ACh as inhibitor, 5HT and FMRFamide as upregulators, with no clear roles for FMRFamide-related peptides and GTP.

Electrical and mechanical responses of the odontophore protractor muscle of Busycon canaliculatum: A sucrose-gap voltage clamp study

Busycon odontophore protractor muscles were examined by conventional force recording and by the single and double sucrose-gap voltage clamp technique. The mean compound resting membrane potential was -61.3 mV and the muscles showed considerable mechanical summation in response to direct electrical stimulation. 2. Responses to high K+ salines consisted of a maintained dose-dependent depolarization upon which were superimposed spike-like action potentials of 3-9 mV each of which generated a fast twitch contraction

Electromechanical uncoupling in a molluscan muscle examined by the sucrose gap technique

SummaryMembrane potential and tension ofBusycon radular protractor muscles were studied by sucrose gap methods.Excitation-contraction (EC) coupling was examined in response to acetylcholine (ACh) and high K which depolarized the fibres and induced tension, but without action potential firing. Potassium depolarization did not follow predictions expected from the Nernst equation at low and very high K levels, and maximum tension was found at about 100 mM K. EC coupling was very sensitive to [Ca]o. Ca-free media eliminated K- and ACh-induced tension but with normal depolarization, showing full electromechanical uncoupling.Ionophore A23187 enhanced K- and ACh-induced responses and X-537A enhanced ACh responses, demonstrating acute dependence of activation on [Ca]o in this muscle. The calcium antagonists nifedipine and nisoldipine reduced tension in the muscle only at very high concentrations, and both agents slightly reduced K- and ACh-induced depolarization.Verapamil reduced K- and ACh-induced tension but paradoxically it enhanced the depolarizing actions of these agents leading to electromechanical uncoupling. Abscisic acid (ABA) enhanced ACh- and K-induced tension and simultaneously enhanced their depolarizing actions. Ionophores and ABA appear to enhance calcium influx which may secondarily influence sodium influx.Calcium antagonists have no consistent actions on this muscle, suggesting that calcium channel activity of the radular protractor may be different from that seen in mammalian visceral muscles.

Close apposition of muscle cells in the longitudinal bands of the body wall of a holothurian, Isostichopus badionotus

SummaryElectron microscopy reveals that sarcolemmata of adjacent muscle cells form pentalaminar junctions by fusion of apposed trilaminar double leaflet membranes. These junctions appear to be candidates for low resistance pathways between muscle fibers. The muscles depolarize slowly when bathed in solutions containing elevated concentrations of KCl, and the sucrose gap method can then be used to measure the potential difference between polarized and depolarized regions. Thus the junctions which we have observed may provide the structural basis for electrical transmission through the sucrose gap.