John D Palmer. American Scientist. Volume 84, Issue 6. Nov/Dec 1996.
I stand at my usual intertidal collecting site near the Marine Biological Laboratory on Cape Cod as the tide recedes, exposing a muddy ocean bottom. At first the area is just a sea of marsh grass, but then suddenly, as if they had arisen by spontaneous generation, literally thousands of fiddler crabs appear. They are so numerous that when I approach them and they flee, I can actually hear their “thunder” as they crash through the grass. During their stay on the surface the crabs are quite active: They feed and fight; the males wave their one large claw in a “come-hither” manner to females who more often than not ignore the enticement.
During the summer months the crabs sit out each high-tide interval in cramped burrows, then all emerge from these grottoes as the ocean above ebbs. The timing of this emergence raises a conundrum to an observer: Even though the tide has fallen and exposed the crabs’ turf, their burrows are still fully flooded-How do they know it is time to emerge? As curious as it may seem, these animals have within their bodies what a romantic like myself calls a living clock, a timepiece that roughly measures off the intermission length between successive tides (an interval of about 12.4 hours).
The study of tidal rhythms began in 1954 when an oxygen-consumption rhythm that appeared to mimic the cycle of the tides was discovered in two species of fiddler crab. Since this pioneering work, tidal rhythms have been found in many denizens of the tidewashed coastlines. Rhythmic fluctuations have been discerned, for example, in locomotion (in crabs), color change (in crabs), swimming (in smaller crustaceans and fish) and shell gaping (in bivalves).
In the past few years, much has been learned about those biological rhythms whose fundamental period matches the solar day. In fruit flies, bread mold and mice, among other organisms, single genes have been shown to be involved in generating these rhythms. Unfortunately, the study of tidal rhythms is not so advanced. In fact, little is known about the genetics of even the most widely studied rhythmic marine animals. When the entire genome of an organism is unknown, locating a specific gene would be more a matter of luck than of skill, and thus the hunt for the genetic basis of tidal rhythms ha not even been attempted.
But the study of tidal clocks lags behind that of solar clocks for a more fundamental reason as well: Unlike the daylength, whose monotonous interval does not deviate from 24 hours thanks to the steady rotation of the earth, the tides are maddeningly changeable. Thus, it is adaptive for living solar-day clocks to be quite accurate-and they are. However, because the ebb and flow of the tide can be quite erratic, there has been little evolutionary pressure for living tidal clocks to be precise-and they are the noisiest known. (The small group of people who study them are some of the most haggard scientists in chronobiology.)
After years of studying biological tidal rhythms, I concluded that some of the difficulty arose from the way the problem had been framed. In 1986, together with my colleague on the other side of the world, Barbara Williams of the Portobello Marine Laboratory in New Zealand, I proposed that typical tide-associated rhythms are not the manifestation of a single clock running at a rate of about 12.4 hours, but two clocks, each running at twice that interval.
If this model is correct, the period of each tidal clock (24.8 hours) differs only slightly from the period of the solar clock (24 hours). Since it is difficult to believe natural selection would be so profligate as to build two clocks that ran at nearly the same rate, I speculate that the same clock that governs solar rhythms also produces tidal rhythms. One felicitous consequence of this proposal would be that the substantial findings already made for solar-day clockworks would then pertain equally to the expression of tidal rhythms.
The crab Sesarma reticulatum lives in the intertidal zone (the strip of shoreline washed by the tides) near the Marine Biological Laboratory in Woods Hole, Massachusetts, where it is exposed to both day/night and tidal cycles. The crab sits out low tides in burrows crowned with chimney-shaped mud entrances and roams the seafloor during high water, especially at night just before midnight. In recognition of its nightly routine, my colleagues and I refer to the animal as the penultimatehour crab.
A microscopic golden-brown alga, the diatom Hantzchia virgata, is also a denizen of intertidal sands near Woods Hole. The tiny diatom can be so abundant that, according to one naturalist, its presence can sometimes be heard as a “gentle sizzling,” presumably the bursting of bubbles of oxygen given off by the tiny plant during photosynthesis. The glassy cell wall of the diatom is perforated by pores, through which it can exude, jet-like, a viscous substance, a process that slowly propels the alga through the sand. By this means the diatom undertakes regular vertical migrations, the inspiration for its common name, the commuter diatom. The diatom burrows into the surface sands and remains there until a daytime low tide exposes its sandflat; it then quickly emerges and piles about five cells deep on the sand surface. Just before the tide returns, it burrows again.
How does the penultimate-hour crab know when it is high tide or the commuter diatom know when it is low tide? The simplest hypothesis would be that the crab or diatom senses a change in an environmental cue, such as alterations in submergence and temperature or pressure changes delivered by the tides. But if either organism is brought into the laboratory and forced to live under conditions in which the major environmental cues are absent (the lights remain constantly on or off, the temperature is unchanging and, of course, there are no tides), its behavior or physiology will still undergo periodic fluctuations. The diatoms, for example, will continue to undertake their vertical migrations on schedule for several weeks. Since these rhythms persist in the absence of environmental cues, the crab and diatom must possess an internal pacemaker, a living clock.
Why would an organism possess an internal clock instead of relying on the environment to keep time? The adaptive significance of a biological clock lies in its ability to alert its owner to periodic changes in the environment in advance of those changes. The commuter diatom has a particularly accurate timepiece, probably a consequence of the unrelenting selection pressure to which it is subjected; any cells that do not burrow in advance of the flood tide are washed away.
This is not to suggest that the period of the living clock is an immutable physical constant like the “period” of a quartz watch, which is set by the resonance frequency of a piezoelectric quartz crystal. Although biological rhythms persist in the constancy of the laboratory, their periods usually change somewhat, becoming slightly longer or shorter than the periods expressed in nature. The rate at which solar-day clocks run in the constancy of the laboratory, for example, tends to be a regular function of the intensity of the artificial light provided. This property has led to use of the prefix circa, which means “about,” in words that designate biological rhythms. A circatidal rhythm is thus a tidal rhythm that in the laboratory has become about one tidal interval in length. In a natural setting, however, the rhythm’s period matches that of the environmental cycle to which the organism is exposed.
This is because the clock is entrained, or conscripted, by periodicities in the environment. The study of many plant and animal rhythms has led chronobiologists to two conclusions about the relation between the living clock and its temporal environment. One is that rhythms are not learned from or impressed on organisms by the environment. Instead they are an expression of the organism’s genome. The other is that the phase of the clock can be altered by environmental cues. The relation between the biological clock and the environment is similar to that between a wristwatch and its owner: The owner cannot make the watch tick faster or slower, but he or she can reset its hands. Solar-day rhythms are reset mainly by the day/night cycles of light, and tidal rhythms are reset by tidal changes in hydrostatic pressure, salinity, mechanical agitation and temperature.
Time and Tide
The fundamental complicating factor in dealing with tidal rhythms is simply the fact that the tides themselves are irregular. On most coastlines of the world there are two high and two low tides each day. The day referred to is not the solar day; it is the lunar day, the 24-hour-and-51-minute average interval between successive moonrises. The tides are produced by gravitational and centrifugal forces generated primarily between the moon and the earth, with some refinements added by the sun.
If one wished to make the best possible predictions of tidal patterns, the four primary drivers would have to be combined with 387 other factors that drive frequency components in the tidal waveform. Typically only the four primary drivers are employed to generate published tide tables. Using one such table I have plotted a month of tidal data for a shoreline near Woods Hole as tide-to-tide deviations from the average tidal period of 12.4 hours, the average interval routinely used to describe the frequency of the tides. As one can see, the tide-to-tide variation in period length is enormous. Moreover, on top of these “scheduled” changes, there are changes to the timing of ebb and flow caused by winds.
Tides vary not only from time to time but also from place to place. Some coastlines in the world experience only one tide per lunar day; the northern margin of the Gulf of Mexico and southeastern Asia are such places. On other coastlines, such as the West Coast of the United States, the tides change on a regular basis from one per day to two. The important point is that the tidal schedule is not perfectly regular: the tides themselves are really circatidal! The 12.4-hour period is an average that can be derived only by combining many days of data.
Because the tides are erratic, selection pressure for the evolution of precise living clocks in intertidal dwellers is weak and varies with the individual and the species under study. The range extends from the rare temporal virtuoso to virtual temporal ciphers. Depending on the species, it is not uncommon to find half a sample in the latter category. Thus most often the chronobiologist who tackles marine organisms has only noisy rhythms to work with.
But noise is not the only factor contributing to his or her befuddlement. The alert reader will have noticed another: Intertidal dwellers are exposed to the day/night cycle as well as to the tide cycle and have been found to display both basic solar-day and tidal periods in a single behavior. The penultimate-hour crab is active during high tides but particularly active during those high tides that occur at night. The commuter diatom, on the other hand, surfaces not during all low tides but only during those low tides that occur during the day.
My experience with Sesarma, the penultimate-hour crab, nicely illustrates the marine chronobiologist’s predicament. The penultimate-hour crab’s rhythms are among the noisiest of the intertidal dwellers, making the species far from ideal for study. Nevertheless, being benighted and easily amused, I embarked on a now-and then four-year study of the activity rhythm of this fractious creature. Of 86 crabs, 20 percent showed only a daily rhythm, 24 percent displayed only a tidal rhythm, 21 percent expressed both daily and tidal components, and the arrhythmic response of the remaining 35 percent made them suitable only for crabcakes.
The Circalunidian Clock
Just as the clock driving solar-day rhythms is thought to have a fundamental (but flexible) 24-hour period that matches that of the rotation of the earth about its axis, the tidal clock had been thought to have a basic (but flexible) 12.4-hour period that matches the average length of the natural tides on most coastlines of the world. Beginning in 1986, however, Barbara Williams and I began to obtain unexpected experimental results that suggested that this intuitive idea might not be correct.
We now postulate that the fundamental period of the clock driving tideassociated rhythms is 24.8 hours, the interval of the lunar day. According to this model, called the circalunidian clock hypothesis, the typical tidal rhythm, which consists of two peaks every day, would be generated by two lunar-day clocks, one driving one peak and the other dock driving the other peak. The two horologes would be tightly coupled 180 degrees out of phase, which would ensure a stable interval of 12.4 hours between peaks. Phase-locked in this manner, the clocks drive rhythms that are easily mistaken for ones governed by a single 12.4-hour clock.
In the past 10 years a great deal of information that supports this hypothesis has accumulated. The pliant-pendulum crab, Helice crassa, lives on the sand and mud flats near the Portobello Marine Laboratory on the South Island of New Zealand. In a large and long study of these crabs, Williams and I found that, brought into the laboratory, most of the crabs displayed the usual, persistent circatidal rhythms. A few of them, however, presented an unanticipated pattern. Alternate peaks in their circatidal rhythm scanned the day at different rates; in other words, they had different circa periods. This independence is inexplicable if the rhythm is the expression of one clock with a constant circatidal period. But it is easily explained by the circalunidian clock hypothesis: The periods differ because the coupling between the two lunidian clocks broke when the crabs were kept in constant conditions, and each clock assumed its own circa period.
Other waveform anomalies also more easily explained by the circalunidian hypothesis are peak vanishing and peak splitting. The activity record of a fiddler crab, Uca pugnax, illustrates vanishing. The crab’s activity rhythm persisted for 14 days in the constant conditions of the laboratory, albeit with a lengthened period. Then, on day 15, alternate activity peaks spontaneously disappeared. The spontaneous disappearance and reappearance of alternate peaks can be interpreted as demonstrating that the peaks are controlled by two clocks, each of which can turn on or off without affecting the other. Peak splitting supports the circalunidian hypothesis for the same reason; the disruption of one of the two intertwined rhythms seems not to affect the other rhythm.
More evidence in support of the circalunidian hypothesis derives from field experiments carried out in Central America, an excellent natural laboratory for marine chronobiology because it offers a choice of oceans. On the Caribbean side of Costa Rica, the shoreline is washed by only one tide each lunar day; fiddler crabs collected there display one activity peak per day in the laboratory. On the Pacific side of Costa Rica, the shore is washed by two tides per lunar day As Franklin Barnwell of the University of Minnesota showed, if Caribbean crabs are transported overland to the Pacific and exposed on that shore for several days, they display two activity peaks per day in the laboratory. Either there is one tidal clock that is able to halve its period, or, as seems more likely, there are two clocks, one of which stops ticking in the absence of environmental cues.
Some fishes also display tidal activity patterns that persist in the constancy of the laboratory. A variation on the fiddler experiment can be done with a common intertidal fish, the shanny, Lipophrysis pholis, which migrates between feeding and resting grounds with each flood tide. When captured and brought into the lab, this fish becomes arrhythmic rather quickly, but rhythmicity is easily restored by exposing caged fish to a few tides. The outcome of exposure depends on the position of the cage on the shore, however. If the cage is inundated by just one tide per day, a unimodal rhythm is initiated, and if the cage is inundated twice a day, the fish displays a bimodal rhythm when it is returned to the laboratory. The rhythm that is established appears to depend on whether one or both of the circalunidian clocks was restarted.
Since its formulation, experimental evidence supporting the circalunidian hypothesis has been obtained from eight species of crabs belonging to five genera, from an isopod (an aquatic version of terrestrial crustaceans such as pill bugs), a snail, a clam and two species of fish. If circalunidian patterning is so ubiquitous, why had it not been observed before? To find such evidence one must either use a large sample or be lucky. This is because the coupling between the two clocks is quite strong and only rarely breaks spontaneously. When the two lunidian clocks are wedded, their combined output is indistinguishable from the typical tidal pattern usually displayed in the laboratory.
Solar and Lunar Clocks?
If the clock that controls tidal rhythms really has a basic period of 24.8 hours, at the very least it must be a near relative to a solar-day clock. Would natural selection have created two separate clocks, whose periods differ by only 3 percent when they are entrained by environmental cues, and overlap greatly when they assume their circa lengths in the laboratory?
This question cannot be settled by reference to common properties of solar-day and lunar-day clocks, such as their common imperviousness to temperature and chemicals. These similarities are not particularly informative because they are adaptive. Evolution would be expected to produce clocks insensitive to temperature and to the chemical environment even if each phylum arose independently by spontaneous generation out of the muck; otherwise the clocks would not run at a constant rate.
It is more revealing to compare the nonadaptive aspects of each clock, which they tend to exhibit only in the artificial conditions of the laboratory. It turns out that solar- and lunar-day clocks fail in the laboratory in similar ways: The rhythms they drive sometimes abruptly assume new periods, disappear and reappear, or split in two. Since there is no reason to think these failure modes have been shaped by evolution, they constitute an argument in favor of a single clock.
Nor is the notion that organisms possess at least two copies of a clock an obstacle to the circalunidian hypothesis. Many animals have been demonstrated to have at least two master clocks, and these clocks sometimes run at different rates. For example, the tenebroid beetle, Blaps gigas, has a clock in each eye that governs sensitivity to light. In the laboratory, these clocks sometimes assume different periods. Such independence is also characteristic of human circadian rhythms; our sleep/wakefulness and body-temperature rhythms sometimes adopt different circa periods in constant conditions and almost always do so for a few days after long jet flights to new time zones.
Indeed some organisms may have more than two copies of the clock. The dinoflagellate, Gonyaulax polyhedra, a unicellular alga that often causes red tides off the California coast, is a fascinating research subject: Not only is the plant motile, it also glows in the dark. It has been studied extensively, particularly in the laboratories of the late Beatrice Sweeney at the Santa Barbara campus of the University of California and of J. Woodland Hastings at Harvard University. Isolated from the sea, the alga continues to undergo rhythms in photosynthesis, cell division, bioluminescent glow, bioluminescent flashing, motility and vertical migration-just to mention a few of the 13 known rhythms. Moreover, when the alga is held under constant conditions, some of these rhythms assume different periods. This raises the possibility that there might be many distinct clocks within this microscopic organism, which is less than 1/20 of a millimeter long.
The only significant problem with the notion that the solar and tidal clocks are one and the same is that day/night cycles, which set the phase of solar-day rhythms, do not affect tidal rhythms. Indeed, it would be nonadapative if they did. If day/night cycles entrained tidal rhythms, the rhythms would no longer be tidal in length. The answer to this conundrum may be that the mechanism that adjusts the clock to environmental cycles is separate from the timekeeping mechanism. Just as a person resets the hands of a watch when he or she arrives in a new time zone, different intermediate machinery may synchronize the clocks to tidal and to solar conditions.
So far I have discussed the biological clock without saying much about its whereabouts and workings. Where are these clocks, and how do they work? Three main approaches have been used in the search for living clocks: exposing an organism to chemicals to see if any will perturb the clock; modifying an organism’s genes to see which ones are involved in rhythmic expression; and extirpating bits and pieces of an organism to find the seat of the clock mechanism. As I have pointed out, one of the happy consequences of the oneclock hypothesis is that work done on the circadian clockworks is germane to marine subjects.
Initial attempts to define the biological clockworks involved subjecting animals and plants to a variety of chemical insults. The idea was to find substances that altered a rhythm’s period or changed its phase, either of which, it was hoped, would signify that the biochemical clockworks had been jostled by the chemical in question. Many substances were tried, including metabolic and respiratory poisons, inhibitors of photosynthesis, mitotic inhibitors and growth factors. Indeed the list of substances that produced no effect grew so long that the more philosophical among us proclaimed that chemical immunity was a necessary precondition for a biological clock: If the clock were sensitive to its chemical environment, it would act as a chemical sensor rather than as a timepiece.
A substance was finally found that consistently lengthened the period of both daily and tidal rhythms: deuterium oxide, or heavy water. Although it was pleasant to succeed after so many failures, experiments with deuterium did not directly illuminate the clockworks. When deuterium is substituted for hydrogen in a cell, almost everything is affected. Compounds built of deuterium are more stable than those built of water, so that reaction rates are changed. It is more viscous than water and therefore slows diffusion rates and decreases ion mobility. And it alters nerve activity and changes the solubility of the respiratory gases. This wide spectrum of effects precludes any specific conclusions about deuterium’s effect on clockworks.
Eventually, however, substances were found that both altered the period or phase of a biological rhythm and targeted specific cellular processes. In fact, a generalization is possible: It appears that a class of compounds that inhibit the assembly of proteins on SOS ribosomes, the ribosomes present on the endoplasmic reticulum and in the cytoplasm of eukaryotic cells, strongly influences biological rhythms. For example, one-hour pulses of the antibiotic anisomycin advance or delay the Gonyaulax bioluminescent-glow rhythm, depending on the phase of the rhythm when the drug is administered.
Protein synthesis is ultimately directed by genetic information coded in nuclear DNA, and, as I mentioned earlier, single genes that affect biological rhythms have now been found in several different organisms, including the fruit fly (Drosophila), the bread mold (Neurospora) and the mouse (Mus). An account of the genetic approach to deciphering the biological clock can be found in “Molecular Biological Clocks,” by Joseph S. Takahashi and Michelle Hoffman, March-April 1995.
In each case, many organisms were subjected to a mutagen and their offspring were then screened for altered daily rhythms. By this means different groups of scientists found the per gene in the fruit fly, the frq gene in the bread mold and the clock gene in the mouse. The mutated genes lengthen or shorten a rhythm’s period, or cause the organism to lose rhythmicity; the clock mutant does both in turn.
I shall use per to describe what is known about these clock genes. The information coded in the gene is periodically translated into messenger ribonucleic acid (mRNA), which is dispatched to the protein-synthesizing machinery in the cell, where the protein itself is assembled. The amounts of per mRNA and protein product present in the cell exhibit circadian rhythms, the mRNA content peaking about seven hours before the protein content. The sequence of amino adds in the per protein identifies it as one of a family of molecules that control the transcription, or copying, of genes from DNA to mRNA, which suggests it might control the expression of its own gene.
DNA sequences similar to those of the wild-type per gene are known in spinach, a green alga called Acetabularia, the chicken, the mouse and humans, suggesting that the gene or its kissing cousins are widely distributed. Unfortunately, as I have mentioned, because little is known about the genetics of intertidal organisms, no one has yet looked for the per-like gene in them.
An Eye on the Time
Where is the clock? The answer depends on the species. In higher vertebrates, including people, it is in two clumps of neurons at the base of the brain called the suprachiasmatic nuclei. In birds, the pineal gland located in the “penthouse” of the brain can act as a clock. And in the cockroach there is a clock in both of the brain’s optic lobes. Although the clockworks may be everywhere similar, its anatomical location is highly various, just as some watchsprings and escapements can be found in wristwatches, wall clocks and pocket watches.
Attempts to locate the seat of the clock in marine organisms are notable for their ingenuity and, given the size of many of these organisms, the dexterity required by the searchers. Neurophysiologists working on vision stumbled on a curious rhythm in two marine mollusks, the sea hare, Aplysia californica, a slow-moving gastropod, and the cloudy-bubble snail, Bulla gouldiana, its near relative. In Jon Jacklet’s laboratory on the Albany campus of the State University of New York, recordings of the intensity of the spontaneous firing of the sea hare’s optic nerve were found to describe a daily waveform. Furthermore, when an eye was excised and kept alive in a solution of essential nutrients, this rhythm persisted unabated!
Each tiny sea-hare eye, less than a millimeter in diameter, consists of a roundish lens, about 4,000 photoreceptor cells and 1,000 secondary neurons. In an attempt to locate the clock within this complex, the eye was surgically whittled down shaving by shaving. It was found that 80 percent of the lens and neurons could be removed without destroying rhythmicity, demonstrating that even part of an isolated nervous system can function as an autonomous clock.
Gene Block’s laboratory at the University of Virginia carried out equally delicate experiments on the eye of the cloudy-bubble snail. In this case the rhythm persisted even when the lens and all of the photoreceptor cells were removed. What was left was the next layer inward, called the basal-retinalneuron stratum. Systematic excision of these cells demonstrated that the rhythm persisted when only six neurons remained. Eventually, when culture and recording techniques were developed that allowed the study of isolated cells, each cell was found to be rhythmic!
The clock in the burrowing amphipod, Corophium volutator, was located by similarly ingenious experimental means. This tiny crustacean sits out low tides in a self-constructed burrow. As the flood tide returns, it emerges, treads water, and allows itself to be swept shoreward to feed. It then rides the ebb tide back to its low-tide residence. This rhythm persists in the laboratory for six days at most, but it can be restarted by subjecting the animal to cold pulses. A pulse of -1.5 degrees Celsius is a sufficient stimulus for summer-collected animals, whereas a chilly degrees is required for those collected in the winter.
This characteristic was exploited in Elfed Morgan’s laboratory at the University of Birmingham to locate Corophium’s clock. The tiny amphipods, which are about the size of a grain of rice, were lashed with thread to rafts made of styrofoam. The raft was then floated on sub-zero ethylene glycol. A thin wire that dangled in the liquid could be bent to touch any part of the constrained animal. The scientists sought to restart the animal’s rhythm by chilling various parts of its anatomy By this means, they discovered that Corophium’s clock is located in its brain.
Similar experiments have been done with the green crab, Carcinus maenas. Like Corophium, the green crab has a tidal activity rhythm that usually does not persist very long in the laboratory but that can be restarted by a cold pulse. Barbara Williams tied arrhythmic crabs to the bottom of an aquarium, which was filled with 15-degree seawater, until only the animal’s eyestalks protruded, like periscopes, above the surface. Then for 10 hours, 1-degree water was dribbled, a drop at a time, onto the eyestalks. This treatment started the tidal rhythm, indicating that important parts of the clock reside in the eyestalks.
Other scientists have found that removing a green crab’s eyestalks not only makes it arrhythmic, it makes it hyperactive. Crude eyestalk extracts injected into hyperactive stalkless crabs cause a Ritalin-like damping of their activity. The inhibition is greatest if the extract is made from a rhythmic crab in the inactive phase of its cycle. The active component of the extracts has been identified as a low-molecular-weight peptide, which has been given the name neurodepressing hormone.
Within the eyestalks, subtending the visual elements, is the X-organ/sinusgland complex, a concatenation of structures that produces and stores various hormones. In Ernest Naylor’s laboratory at the University of North Wales, holes were drilled through intact eyestalks and specific parts of the innards destroyed by cautery Only when the Xorgan was incinerated did rhythmic locomotion cease. In another experiment, staining X-organ cells with a dye revealed that during the inactive phase of the crabs’ rhythm neurosecretory material was produced primarily by one cell type. Attempts to demonstrate rhythmicity in isolated eyestalk tissues have so far failed, however, and so the selfcontainment of the X-organ clock must be regarded as unproved.
A Timely Conundrum
For the sake of those readers still not suitably impressed by the difficulty (and charm) of marine chronobiology, I conclude with a horological conundrum, that posed by the unicellular green alga, Acetabularia. Depending on the species, a single cell of this filamentous green alga can reach an astonishing length of 13 centimeters. The alga expresses several persistent rhythms, but here I focus on its photosynthetic rhythm. In the early stages of its life cycle Acetabularia consists of a root-like holdfast that anchors it to the substratum as an upright green filament. Each cell undergoes a daily cycle in photosynthetic capacity, which peaks during daytime.
The nucleus, the master control center, resides in the holdfast at this stage of development. If the nucleus is removed by cutting off the holdfast, the enucleated alga will live for up to three months. But that feat is just a petty prelude to what follows: Not only will the photosynthetic rhythm persist in constant conditions, it will do so for as long as a month after the nucleus has been removed. Thus this organism demonstrates unambiguously that not only are endocrine and nervous systems not needed for rhythmicity, neither is a nucleus. The last trick I shall describe is even more spectacular: If a single filament is cut into 10-millimeter fragments, each tiny amputee continues to be rhythmic. Apparently, there are many clock replicates in each Acetabularia cell, or as John Aldrich of Trinity College puts it, in this case the anatomical location of the clock seems to be “just about everywhere.”