Anthony A. Paparo

RAPID ASSESSMENT OF WATER QUALITY, USING THE FINGERNAIL CLAM, MUSCULIUM TRANSVERSUM

An apparatus for testing the effects of drugs on the ciliary beating rate of clam gills has been modified to rapidly (15 min to 1 h) assess the effects of water quality factors on a sensitive organism, the fingernail clam, Musculium transversum. The gill and adductor muscles of the clam are excised and placed in a petri dish through which a continuous flow of molluscan Ringers solution or a test solution can be maintained. Normal ciliary activity of the gill preparation can be maintained for at least eight days. The ciliary beating rate is determined by synchronizing the rate of flashing of the sub-stage lamp of a microscope, with the rate of beating of the cilia. Synchronization is achieved when the metachronal ciliary wave appears to stand still. The first water quality factor selected for testing by the rapid method was potassium, because potassium concentrations are higher in the Illinois River where fingernail clams have largely died out, than in the Mississippi River where the clams are still abundant. The apparatus provided statistically reliable results in a short period of time. There are significant differences in the responses of large (7 to 11 mm) and small (1 to 5 mm) clams to: (a) removal and subsequent addition of potassium, (b) variation of maintenance dosage of potassium in the washing solution, and (c) lag period of response to a specific dose. The results suggest that an intracellular trans-membrane potential change (surface effect) is necessary to activate ciliated cells of small clams. This latter change in small clams would account for the relatively short lag period for potassium activation. Potassium levels required for maintenance of a basal ciliary beating rate are 10 - 3 M (39.1 mg/litre) for small clams and 10 - 6 M (0.039 mg/litre) for large clams. Greater concentrations are cilioinhibitory. Lesser concentrations are generally cilioexcitatory, but concentrations less than 10 - 8 M (0.00039 mg/litre) and 10 - 9 M (0.000039 mg/litre) are insufficient to sustain basal rates in large and small clams, respectively. Potassium concentrations in certain rivers, such as the Illinois and Mississippi, are high enough to cause cilioinhibition in gill preparations from large fingernail clams. Inhibition of ciliary activity in intact clams would impair feeding and respiration. The effects of potassium on the survival, growth, and reproduction of intact fingernail clams are currently being determined and will be related to the effects observed by means of the rapid method.

The regulation of intracellular calcium and the release of neurotransmitters in the mussel, Mytilus edulis

1. 1. Lateral ciliary activity was studied on isolated visceral ganglion/branchial nerve/gill preparations of Mytilus edulis. 2. 2. Branchial nerve stimulation with 5 pulses per sec and perfusion with serotonin each accelerated ciliary beating. 3. 3. Branchial nerve stimulation with 50 pulses per sec and perfusion with dopamine each inhibited ciliary activity. 4. 4. A decrease in external free calcium ion concentration or interference with calcium membrane permeability blocks nerve stimulation (low and high frequency) and subsequent endogenous release of neurotransmitters. Exogenous application of serotonin and dopamine is not blocked by manipulation of calcium concentrations or calcium membrane interactions. 5. 5. Calcium plays a critical role in the regulation of presynaptic nerve terminal release of serotonin and dopamine.

The effect of various salinities on whole and dissected preparations of Crassostrea virginica

Abstract 1. 1. All experiments were performed on whole animal or isolated preparations of the American oyster, Crassostrea virginica . 2. 2. Whole preparations showed a linear decline in the transport of particles when the salinities were changed. Dissected preparations exhibited a transport rate which declined more precipitously. 3. 3. The dissected preparation demonstrated a decrease in serotonin and DOPA decarboxylase activities. This was further exacerbated in the presence of PTZ (an intracellular calcium releasor) and salyrgan (a calcium-pump inactivator) perfusates. 4. 4. These experiments support the hypothesis that the effect of salinity change on transport of particles is in fact an effect of the change in calcium ion concentration.

The relationship between lamellar-type cytosomes, DOPA decarboxylase, and lateral ciliary activity of the oyster Crassostrea virginica (Gmelin): Response to salinity change

Abstract Lamellar transformations of cytosomes, DOPA decarboxylase and lateral ciliary activity were measured in neuronal components and/or ctenidia of the oyster, Crassostrea virginica (Gmelin). Prolonged exposure to light, anoxic conditions, EGTA (a calcium chelator), and exogenous perfusion of dopamine (a cilioinhibitory neurotransmitter) increases the frequency of lamellar type cytosomes. This is accompanied by decreases in enzymatic activity and a lateral cilioinhibition. Prolonged periods of darkness, electrical stimulation of the branchial nerve and exogenous perfusion of serotonin (a cilioexcitatory neurotransmitter) reduces the conversion of lamellar type configurations, increases enzymatic activity and produces a lateral cilioexcitation. Salinity changes that produce an increase to higher acclimation values result in a gradual decrease in the number of lamellar configurations with a subsequent increase in enzymatic activity and lateral cilioexcitation. The magnitude of the lateral cilioexcitation-cilioinhibition and decarboxylase activity is directly related to the number of lamellar conversions.

The role of the cerebral and visceral ganglia in ciliary activity

Abstract 1. 1. Lateral ciliary activity was studiedon isolated gill preparations of the mussel Mytilus edidis. 2. 2. Electrical stimulation of the branchial nerve at 5 Hz (increased) and 50 Hz (decreased) changed the average rate of ciliary beating. 3. 3. These modifications in the ciliary activity were significantly altered in specimens lacking an intact cerebrovisceral connective. 4. 4. It appears that the cerebral and visceral ganglia exhibit a coordinated role in the control of ciliary beating.

A role for acetylcholine in the regulation of ciliary activity

Abstract 1. Low concentrations of acetylcholine and physostigmine were excitatory and high concentrations were inhibitory to lateral cilia of Mytilus edulis gill. 2. Physostigmine potentiated the effects of acetylcholine and nerve stimulation. 3. Bromolysergic acid diethylamide blocked excitation by acetylcholine, physostigmine, 5-hydroxytryptamine and 5 Hz nerve stimulation. 4. Phenoxybenzamine blocked inhibition by acetylcholine, physostigmine, dopamine and 50 Hz nerve stimulation. 5. It is postulated that acetylcholine affects ciliary beating by releasing endogenous mediators and that this is the principal physiological role of endogenous acetylcholine regarding ciliary activity in the gill.

DOPA decarboxylase from visceral ganglion and gill of two lamellibranch molluscs as it relates to ciliary activity

Abstract 1. DOPA decarboxylase was measured in the visceral ganglion and gill epithelium of Mytilus edulis and Elliptio complanata. 2. DOPA decarboxylase inhibitors (STH, HMD, and α-methyl-DOPA) reduced decarboxylase activity in extracts of visceral ganglion and gill in both species. 3. DOPA decarbroxylase activity was decreased by ligating or severing the branchial nerve in gill preparations. 4. 5-HTP decarboxylase in the visceral ganglion is about 15–20 per cent of DOPA decarboxylase activity. 5. Intramuscular injection of STH, HMD causes the rate of beating of the lateral cilia to become independent of stimulation. 6. The cilio-excitatory effect of branchial nerve stimulation in vitro decreased after ligation of the branchial nerve and intramuscular injection of 6-OH-DOPA and of DOPA decarboxylase inhibitors. 7. Therefore, it appears that ciliary activity in the gill is dependent upon the decarboxylase activity of the visceral ganglion and branchial nerve.

Ciliary activity on the ctenidium of bivalve molluscs

Abstract 1. Lateral ciliary activity was studied on the ctenidial preparations of several bivalves. 2. The cerebral and visceral ganglia exhibit a coordinated role in the control of ciliary beating. 3. Exposure of the ctenidia to changes in potassium and magnesium ion concentrations at acclimation salinities were salinity-dependent and probably reflect an effect on the ciliated epithelium. 4. The magnitude of cilio-inhibition is directly related to the percentage of lamellar conversions in cytosomes. 5. Since sequestered calcium has been shown to be released from lamellar-type cytosomes, it is postulated that lateral cilio-inhibition is due to an increase in the neuronal intracellular calcium concentration with subsequent release of the cilio-inhibitory neurotransmitter, dopamine. 6. There is a seasonal effect on cytosomal transformations and decarboxylase activity in neuronal tissues both in the central and peripheral elements and lateral ciliary activity. 7. The experimental design and procedures used in our studies will have broad applications for quantitatively assessing the effects of environmental factors on ciliary activity of marine and estuarine organisms.

Average ciliary beat in the oyster: response to photoperiod, pentylenetetrazole, salyrgan, serotonin and dopamine

Lamellar conversions of cytosomes, DOPA decarboxylase and lateral ciliary activities were measured in central and peripheral neuronal tissues and/on ctenidia of the oyster, Crassostrea virginica (Gemelin). 5‰ S habitat and acclimation animals exhibited significantly (P < 0.0S) higher numbers of transformed cytosomes and lower decarboxylase activities. Exposure to light for long periods of time, and exogenous perfusion at pentylenetetrazole enhanced lamellar conversion. Periods of darkness reduced the incidence of lamellar morphogenesis. Salyrgan can both increase (in 5‰ S and 10‰ S groups) and decrease (in 15‰ through 30‰ S groups) membranous conversions. Electrical stimulation of the branchial nerve increased the average lateral ciliary beat and decarboxylase activities in the ctenidium. Serotonin (cilioex‐citatory) and dopamine (cilioinhibitory) are endogenous neurotransmitters. The magnitude of the lateral cilioexcitation‐cilioinhibition and decarboxylase activity is directed related to the incidence o...

Activity of the lateral cilia of the oyster Crassostrea virginica gmelin: Response to changes in salinity and to changes in potassium and magnesium concentrationt†

Lateral ciliary activity (LCA) on ctenidial filaments of the oyster has been measured in animals acclimated to 30, 15 and 5%o seawater from three to five days. Using shucked, intact tissue with the minimum dissection required to expose the ctenidia, the responses of LCA to rapid changes in salinity have been defined. A linear decline in LCA was observed over the 140 minute exposure period; the slope of the regression line and the percent inhibition of the activity were both proportional to the magnitude of salinity change and were useful indices to the effect of the test medium. The effect of exposure of tissues to changes in potassium ion concentration at the acclimation salinities was found to be salinity‐dependent and is probably exerted on the branchial nerve rather than the ciliated epithelium. Exposure of tissues to changes in magnesium ion concentrations at the acclimation salinities was also salinity‐dependent and probably reflects an effect directly on the ciliated epithelium. The experimental de...