Rhythmic Output Research Articles

Abnormal Diurnal Rhythm of Urine Output Following Renal Transplantation: The Impact of Blood Pressure and Diuretics

Background Nocturnal polyuria is the excretion at night of an excessive volume of urine. A major problem following renal transplantion is an abnormal diurnal rhythmicity in urine output. The purpose of this study was to elucidate the prevalence of nocturnal polyuria among renal transplant recipients in the early period after transplantation as well as at least 1 year after transplantation. We aimed to explore possible pathophysiological mechanisms behind nocturnal polyuria in this group of patients, focusing on the impact of blood pressure and medication. Methods Seventeen recently transplanted patients 17 late transplant recipients, and 17 healthy controls were included in the study. Voiding habits were assessed by completion of a frequency-volume chart recording all fluid intakes and voiding. A concomitant 24-hour blood pressure profile was obtained in all. Results Renal transplant recipients had a high prevalence of nocturnal polyuria (74%) and a disturbed blood pressure profile with a lack of appropriate nocturnal dipping ( P < .0001) compared to controls. We found a positive correlation between increased nocturnal blood pressure and urine output at night ( r = .368, P = .008). Patients taking diuretics had a circadian diurnal rhythm of urine output and a blood pressure profile similar to controls. Conclusions Nocturnal polyuria was very common among both recent and late transplant recipients. A high fluid intake during daytime and hypervolemia were suggested as causes of a disturbed blood pressure profile, which partly seemed to explain the high urine output at night. Daytime diuretics may be an effective treatment of this inconvenient complication.

Processing Circadian Data Collected from the Drosophila Activity Monitoring (DAM) System: Figure 1.

Adult behavioral assays have been used with great success in Drosophila melanogaster to identify circadian rhythm genes. In particular, the locomotor activity assay can identify altered behavior patterns over the course of several days in small populations, or even individual flies. Generally, circadian behavior is assayed during a period of 12 h light:12 h dark cycling (LD entrainment) followed by conditions of constant darkness (DD). LD activity profiles provide a qualitative image of daily activity bouts, and the data can be used to quantitatively assess the phase and/or amplitude of particular bouts. Additional activity assessments made from entrained flies that have been shifted to constant darkness can provide insight into the state of internal clocks and the ability of these clocks to drive rhythmic outputs. Typical LD DD runs assess both free-running rhythmicity and period length with χ2 periodogram (P'gram) analysis. This protocol describes the use of ClockLab (a MATLAB-based program) and the Counting Macro (an Excel-based program) to analyze circadian locomotor activity data collected using the Drosophila Activity Monitoring (DAM) System from TriKinetics. Specific procedures are described to analyze free-running rhythmicity and period length for individual flies, and assess group activity plots during both entrainment and constant conditions.

Central pattern generation underlying Limulus rhythmic behavior patterns

Abstract Many behavioral activities of the horseshoe crab Limulus are rhythmic, and most of these are produced in large part by central pattern generators within the CNS. The chain of opisthosomal ('abdominal') ganglia controls gill movements of ventilation and gill cleaning, and the prosomal ring of fused ganglia (brain and segmental 'thoracic' ganglia) controls generation of feeding and locomotor movements of the legs. Both the opisthosomal CNS and the prosomal CNS can generate behaviorally appropriate patterns of motor output in isolation, without movements or sensory input. Preparations of the isolated opisthosomal CNS generate rhythmic output patterns of motor activity characterized as fictive ventilatory and gill cleaning rhythms. Moreover, CNS preparations also express longer-term patterns, such as intermittent ventilation or sequential bouts of ventilation and gill cleaning. Such longer-term patterns are commonly observed in intact animals. The isolated prosomal CNS does not spontaneously generate the activity patterns characteristic of walking, swimming, and feeding. However, perfusion of octopamine in the isolated prosomal CNS activates central pattern generators underlying rhythmic chewing movements, and injection of octopamine into intact Limulus promotes the chewing pattern of feeding, whether or not food is presented. Our understanding of the ability of neuromodulators such as octopamine to elicit or alter central motor programs may help to clarify the central neural circuits of pattern generation that produce and coordinate these rhythmic behaviors.

Review: Clock genes at the heart of depression

The rhythms of life are ever pervasive, touching almost every aspect of our lives. We are finely tuned to the cycle of light and dark, so that we normally sleep during the night and are active during the day. Physiological rhythms are, however, not just slaves to the solar day, but are actually generated endogenously within the suprachiasmatic nuclei in the hypothalamus and are entrained via the retina. The circadian timing system is organized hierarchically with the suprachiasmatic nuclei providing neural and/or hormonal cues to the various organ systems, allowing them to express their own rhythmic physiological output. There is now a substantial body of evidence emerging that disruption of rhythmicity through altered sleep/wake patterns and exposure to light, or through endogenous disruption of key determinants of endogenous rhythms, can be detrimental to health. Circadian rhythm disturbances have long been associated with mood disorders, especially delayed sleep onset, and evidence is accumulating that alterations to the cellular timing system may underpin some aspects of the disorders. For example, mice carrying mutations in either Clock or per2 spend less time immobile in swim tests, which has been suggested as mimicking mania. In humans, single nucleotide polymorphisms in Clock and other clock genes have been associated with depression. With this increasing knowledge we may predict that new antidepressant drugs will emerge that, as a primary or secondary mechanism of action, target and correct abnormalities in the circadian timing system.

Correct biological timing inArabidopsisrequires multiple light-signaling pathways

Circadian oscillators provide rhythmic temporal cues for a range of biological processes in plants and animals, enabling anticipation of the day/night cycle and enhancing fitness-associated traits. We have used engineering models to understand the control principles of a plant's response to seasonal variation. We show that the seasonal changes in the timing of circadian outputs require light regulation via feed-forward loops, combining rapid light-signaling pathways with entrained circadian oscillators. Linear time-invariant models of circadian rhythms were computed for 3,503 circadian-regulated genes and for the concentration of cytosolic-free calcium to quantify the magnitude and timing of regulation by circadian oscillators and light-signaling pathways. Bioinformatic and experimental analysis show that rapid light-induced regulation of circadian outputs is associated with seasonal rephasing of the output rhythm. We identify that external coincidence is required for rephasing of multiple output rhythms, and is therefore important in general phase control in addition to specific photoperiod-dependent processes such as flowering and hypocotyl elongation. Our findings uncover a fundamental design principle of circadian regulation, and identify the importance of rapid light-signaling pathways in temporal control.

Changes in electrical coupling via dynamic clamp produces correlates of operant conditioning in the feeding CPG networks of Aplysia

Appetitive operant conditioning, a form of associative learning, produces a long-lasting switch in the mollusk Aplysia’s food-seeking behavior from irregular, impulsive-like radula biting movements into stereotyped, compulsive-like recurrences of this cyclic act [1]. Three bilateral pairs of neurons (B63, B65, and B30) in the feeding central pattern generator (CPG) circuit, found in the buccal ganglia, are initiators of each radula bite motor pattern. Using isolated ganglia from naive and operantly conditioned animals, it was previously found that learning increased both the frequency and regularity of spontaneous bursting activity in these three pairs of neurons. This plasticity was correlated with an increase in electrical coupling among these cells, together with an increased excitability and a change in their intrinsic oscillatory membrane properties [2]. In the present study, we explored the role that changes in electrical coupling between the 6 neurons might play in the operant learning-induced regularization and acceleration of biting motor pattern genesis. In isolated buccal ganglia, we implemented a dynamic clamp procedure during fictive biting activity to artificially increase the electrical coupling among the identified motor pattern initiating neurons. In preparations which spontaneously generated irregular biting motor patterns, artificially increasing the coupling among groups of any 3-4 (out of the 6) neurons immediately switched the network from an irregular to a regular burst pattern. Conversely, in preparations in which the motor pattern-initiating cells spontaneously generated rhythmic motor output, a decrease in their electrical coupling switched the cell’s bursting activity and radula motor pattern genesis from a rhythmic to an arrhythmic mode. These changes were not correlated to a change either in resting membrane potential of the recorded cells or in the frequency of their impulse bursts. These data therefore suggest a causal relationship between the strength of electrical coupling and the temporal regularity of motor pattern genesis underlying each radula cycle, while a separate mechanism is likely to account for the changes in motor pattern frequency. Plasticity induced by operant learning in electrical connectivity within a subset of CPG neurons, may thereby provide a cellular substrate for the behavioral switch between impulsive to compulsive motor actions in Aplysia as well as in more complex animals.

Vasopressin and the Output of the Hypothalamic Biological Clock

The physiological effects of vasopressin as a peripheral hormone were first reported more than 100 years ago. However, it was not until the first immunocytochemical studies were carried out in the early 1970s, using vasopressin antibodies, and the discovery of an extensive distribution of vasopressin-containing fibres outside the hypothalamus, that a neurotransmitter role for vasopressin could be hypothesised. These studies revealed four additional vasopressin systems next to the classical magnocellular vasopressin system in the paraventricular and supraoptic nuclei: a sexually dimorphic system originating from the bed nucleus of the stria terminalis and the medial amygdala, an autonomic and endocrine system originating from the medial part of the paraventricular nucleus, and the circadian system originating from the hypothalamic suprachiasmatic nuclei (SCN). At about the same time as the discovery of the neurotransmitter function of vasopressin, it also became clear that the SCN contain the main component of the mammalian biological clock system (i.e. the endogenous pacemaker). This review will concentrate on the significance of the vasopressin neurones in the SCN for the functional output of the biological clock that is contained within it. The vasopressin-containing subpopulation is a characteristic feature of the SCN in many species, including humans. The activity of the vasopressin neurones in the SCN shows a pronounced daily variation in its activity that has also been demonstrated in human post-mortem brains. Animal experiments show an important role for SCN-derived vasopressin in the control of neuroendocrine day/night rhythms such as that of the hypothalamic-pituitary-adrenal and hypothalamic-pituitary-gonadal axes. The remarkable correlation between a diminished presence of vasopressin in the SCN and a deterioration of sleep-wake rhythms during ageing and depression make it likely that, also in humans, the vasopressin neurones contribute considerably to the rhythmic output of the SCN.

Impact of aging on diurnal expression patterns of CLOCK and BMAL1 in the mouse brain

Mammalian circadian rhythms are generated by a network of transcriptional and translational loops in the expression of a panel of clock genes in various brain and peripheral sites. Many of the output rhythms controlled by this system are significantly affected by ageing, although the mechanisms of age-related circadian dysfunction remain opaque. The aim of this study was to investigate the effect of aging on the daily oscillation of two clock gene proteins (CLOCK, BMAL1) in the mouse brain. Clock gene protein expression in the brain was measured by means of immunohistochemistry in groups of young (4 months) and older (16 months) mice sampled every 4 h over a 24-h cycle. CLOCK and BMAL1 were constitutively expressed in the suprachiasmatic nucleus (SCN; the master circadian pacemaker) in young adult animals. We report novel rhythmic expression of CLOCK and BMAL1 in a number of extra-SCN sites in the young mouse brain, including the hippocampus, amygdala and the paraventricular, arcuate and dorsomedial nuclei of the hypothalamus. Aging altered the amplitude and/or phase of expression in these regions. These results indicate hitherto unreported expression patterns of CLOCK and BMAL1 in non-SCN brain circadian oscillators, and suggest that alterations of these patterns may contribute to age-related circadian dysfunction.

Rhythmic behaviour and pattern-generating circuits in the locust: Key concepts and recent updates

There is growing recognition that rhythmic activity patterns are widespread in our brain and play an important role in all aspects of the functioning of our nervous system, from sensory integration to central processing and motor control. The study of the unique properties that enable central circuits to generate their rhythmic output in the absence of any patterned, sensory or descending, inputs, has been very rewarding in the relatively simple invertebrate preparations. The locust, specifically, is a remarkable example of an organism in which central pattern generator (CPG) networks have been suggested and studied in practically all aspects of their behaviour. Here we present an updated overview of the various rhythmic behaviours in the locust and aspects of their neural control. We focus on the fundamental concepts of multifunctional neuronal circuits, neural centre interactions and neuromodulation of CPG networks. We are certain that the very broad and solid knowledge base of locust rhythmic behaviour and pattern-generating circuits will continue to expand and further contribute to our understanding of the principles behind the functioning of the nervous system and, indeed, the brain.

The brain can eat: Establishing the existence of a central pattern generator for feeding in third instar larvae of Drosophila virilis and Drosophila melanogaster

To establish the existence of a central pattern generator for feeding in the larval central nervous system of two Drosophila species, the gross anatomy of feeding related muscles and their innervation is described, the motor units of the muscles identified and rhythmic motor output recorded from the isolated CNS. The cibarial dilator muscles that mediate food ingestion are innervated by the frontal nerve. Their motor pathway projects from the brain through the antennal nerves, the frontal connectives and the frontal nerve junction. The mouth hook elevator and depressor system is innervated by side branches of the maxillary nerve. The motor units of the two muscle groups differ in amplitude: the elevator is always activated by a small unit, the depressor by a large one. The dorsal protractors span the cephalopharyngeal skeleton and the body wall hence mediating an extension of the CPS. These muscles are innervated by the prothoracic accessory nerve. Rhythmic motor output produced by the isolated central nervous system can simultaneously be recorded from all three nerves. The temporal pattern of the identified motor units resembles the sequence of muscle contractions deduced from natural feeding behavior and is therefore considered as fictive feeding. Phase diagrams show an almost identical fictive feeding pattern is in both species.

Arrhythmic Rats after SCN Lesions and Constant Light Differ in Short Time Scale Regulation of Locomotor Activity

Circadian rhythm disruption (i.e., arrhythmicity) in motor activity is an abnormal behavioral pattern. In rats, it can be caused by the lesion of the hypothalamic suprachiasmatic nucleus (SCN) and by prolonged exposure to constant light (LL). We carried out a comparative study of these arrhythmic phenotypes to assess the role of the SCN in the regulation of the motor output beyond circadian rhythmicity. Motor activity series were studied in rats that had become arrhythmic as a result of 1) LL exposure at 2 light intensities: 300 lux (LL(300)) and 1.3 lux (LL(1.3)), and 2) SCN lesion (SCNx). The Fourier spectra, the fractal Hurst coefficient (H) from the autocorrelation function, and the beta slope from the power spectral density were calculated in data sections at baseline, when the rats were still rhythmic, and later at stages with undetectable circadian rhythms. In the LL(300) group, high power content was detected at frequencies of 8 to 4 h (i.e., ultradian). Lower power content for these harmonics was found in the LL(1.3) group, whereas no dominant harmonics appeared in the SCNx group. Independently of the manifestation of circadian rhythm, H values were higher and more sustained in time in rats exposed to LL( 300) but gradually decreased in rats exposed to LL(1.3). Fractal correlation was found in control DD group but was absent in the SCNx group. We conclude that scale-invariant regulation of the motor pattern by SCN activity is dependent on light intensity but independent of the circadian rhythm output. Adjusting the light intensity by modifying the coupling degree between the population of oscillations could affect the dynamics of each individual oscillator in the SCN, making it less predictable.

Postembryonic development of centrally generated flight motor patterns in the hawkmoth, Manduca sexta

This study analyses the maturation of centrally generated flight motor patterns during metamorphosis of Manduca sexta. Bath application of the octopamine agonist chlordimeform to the isolated central nervous system of adult moths reliably induces fictive flight patterns in wing depressor and elevator motoneurons. Pattern maturation is investigated by chlordimeform application at different developmental stages. Chlordimeform also induces motor patterns in larval ganglia, which differ from fictive flight, indicating that in larvae and adults, octopamine affects different networks. First changes in motoneuron activity occur at the pupal stage P10. Rhythmic motor output is induced in depressor, but not in elevator motoneurons at P12. Adult-like fictive flight activity in motoneurons is observed at P16 and increases in speed and precision until emergence 2 days later. Pharmacological block of chloride channels with picrotoxin also induces fictive flight in adults, suggesting that the pattern-generating network can be activated by the removal of inhibition, and that proper network function does not rely on GABA(A) receptors. Our results suggest that the flight pattern-generating network becomes gradually established between P12 and P16, and is further refined until adulthood. These findings are discussed in the context of known physiological and structural CNS development during Manduca metamorphosis.

Math1 Is Essential for the Development of Hindbrain Neurons Critical for Perinatal Breathing

Mice lacking the proneural transcription factor Math1 (Atoh1) lack multiple neurons of the proprioceptive and arousal systems and die shortly after birth from an apparent inability to initiate respiration. We sought to determine whether Math1 was necessary for the development of hindbrain nuclei involved in respiratory rhythm generation, such as the parafacial respiratory group/retrotrapezoid nucleus (pFRG/RTN), defects in which are associated with congenital central hypoventilation syndrome (CCHS). We generated a Math1-GFP fusion allele to trace the development of Math1-expressing pFRG/RTN and paratrigeminal neurons and found that loss of Math1 did indeed disrupt their migration and differentiation. We also identified Math1-dependent neurons and their projections near the pre-Bötzinger complex, a structure critical for respiratory rhythmogenesis, and found that glutamatergic modulation reestablished a rhythm in the absence of Math1. This study identifies Math1-dependent neurons that are critical for perinatal breathing that may link proprioception and arousal with respiration.

Dissociation of circadian and light inhibition of melatonin release through forced desynchronization in the rat

Pineal melatonin release exhibits a circadian rhythm with a tight nocturnal pattern. Melatonin synthesis is regulated by the master circadian clock within the hypothalamic suprachiasmatic nucleus (SCN) and is also directly inhibited by light. The SCN is necessary for both circadian regulation and light inhibition of melatonin synthesis and thus it has been difficult to isolate these two regulatory limbs to define the output pathways by which the SCN conveys circadian and light phase information to the pineal. A 22-h light-dark (LD) cycle forced desynchrony protocol leads to the stable dissociation of rhythmic clock gene expression within the ventrolateral SCN (vlSCN) and the dorsomedial SCN (dmSCN). In the present study, we have used this protocol to assess the pattern of melatonin release under forced desynchronization of these SCN subregions. In light of our reported patterns of clock gene expression in the forced desynchronized rat, we propose that the vlSCN oscillator entrains to the 22-h LD cycle whereas the dmSCN shows relative coordination to the light-entrained vlSCN, and that this dual-oscillator configuration accounts for the pattern of melatonin release. We present a simple mathematical model in which the relative coordination of a single oscillator within the dmSCN to a single light-entrained oscillator within the vlSCN faithfully portrays the circadian phase, duration and amplitude of melatonin release under forced desynchronization. Our results underscore the importance of the SCN's subregional organization to both photic input processing and rhythmic output control.

Descending interactions with spinal cord networks: a time to build?

Descending interactions with spinal cord networks: a time to build?

Limits of single-cell autonomy in the suprachiasmatic nucleus

The suprachiasmatic nucleus (SCN) is the master circadian pacemaker in the mammalian brain, responsible for coordinating circadian (ca 24 h) rhythms throughout the body. When individual SCN neurons are dispersed in low density culture, they generate independent circadian oscillations in neuronal firing rate. This observation has led to the widely accepted conclusion that single SCN neurons are autonomous circadian oscillators. More recent studies using bioluminescence imaging to monitor rhythms of clock gene expression have indicated that fibroblasts, too, are autonomous circadian oscillators. But does this mean that the fibroblast should “replace the SCN as the in vitro model of choice”? On the contrary, the SCN is much more than a population of independent cellular oscillators. SCN function depends on important network interactions that allow for cell synchrony, enhanced pacemaker precision, reinforcement of rhythm amplitude, and robustness against genetic perturbations. The distribution of phases of cells within the SCN network also permits variously phased rhythmic output signals, as well as the encoding of seasonal changes in photoperiod and other features of environmental lighting history. Thus, unlike fibroblasts, SCN neurons are designed to be “team players,” and SCN network interactions are integral to the normal function of the SCN as a circadian pacemaker.

Analysis of theDrosophila ClockPromoter Reveals Heterogeneity in Expression between Subgroups of Central Oscillator Cells and Identifies a Novel Enhancer Region

The CLOCK-CYCLE (CLK-CYC) heterodimer lies at the heart of the circadian oscillator mechanism in Drosophila, yet little is known about the identity of transcription factors that regulate the expression of Clk and/or cyc. Here, the authors have used a transgenic approach to isolate regions of the Clk locus that are necessary for expression in central oscillator neurons in the adult fly brain. This analysis shows that central clock cells can be subdivided into 2 distinct groups based on Clk gene regulation. Expression in the lateral neuron (LN), dorsal neuron 1 anterior (DN1a) and 2 (DN2) clusters requires cis-elements located in a 122 base-pair (bp) region (-206 to -84) of the Clk promoter. Expression in the remaining dorsal neurons, 1 posterior (DN1p) and 3 (DN3) and the lateral posterior neurons (LPN), requires regulatory elements located in the -856 to -206 region. In addition, expression in photoreceptors of the compound eye is enhanced by cis-elements located in a 3rd region of the Clk locus (-1982 to -856). This region also enhances expression in nonoscillator cells in the brain including the Kenyon cells, but expression in these neurons is suppressed by regulatory sites located further upstream of -1982. The authors' analysis reveals clear heterogeneity in Clk gene expression in the adult brain and provides a necessary focus to isolate novel transcription factors that bind at the Clk locus to regulate expression in different oscillator neuron subgroups. These results also suggest that the DN1a/DN2 neurons may have more molecular commonality with the LNs than they do with the DN1p/DN3/LPN neurons. Finally, this analysis has generated new transgenic lines that will enable genes to be misexpressed in subgroups of central oscillator cells that have previously been resistant to discrete genetic manipulation. Hence, these lines provide important new tools to facilitate a more complete dissection of the neural network that regulates output rhythms in physiology and behavior.

Function of the Shaw potassium channel within the Drosophila circadian clock

Background In addition to the molecular feedback loops, electrical activity has been shown to be important for the function of networks of clock neurons in generating rhythmic behavior. Most studies have used over-expression of foreign channels or pharmacological manipulations that alter membrane excitability. In order to determine the cellular mechanisms that regulate resting membrane potential (RMP) in the native clock of Drosophila we modulated the function of Shaw, a widely expressed neuronal potassium (K+) channel known to regulate RMP in Drosophila central neurons. Methodology/Principal Findings We show that Shaw is endogenously expressed in clock neurons. Differential use of clock gene promoters was employed to express a range of transgenes that either increase or decrease Shaw function in different clusters of clock neurons. Under LD conditions, increasing Shaw levels in all clock neurons (LNv, LNd, DN1, DN2 and DN3), or in subsets of clock neurons (LNd and DNs or DNs alone) increases locomotor activity at night. In free-running conditions these manipulations result in arrhythmic locomotor activity without disruption of the molecular clock. Reducing Shaw in the DN alone caused a dramatic lengthening of the behavioral period. Changing Shaw levels in all clock neurons also disrupts the rhythmic accumulation and levels of Pigment Dispersing Factor (PDF) in the dorsal projections of LNv neurons. However, changing Shaw levels solely in LNv neurons had little effect on locomotor activity or rhythmic accumulation of PDF. Conclusions/Significance Based on our results it is likely that Shaw modulates pacemaker and output neuronal electrical activity that controls circadian locomotor behavior by affecting rhythmic release of PDF. The results support an important role of the DN clock neurons in Shaw-mediated control of circadian behavior. In conclusion, we have demonstrated a central role of Shaw for coordinated and rhythmic output from clock neurons.

The Circadian System in Higher Plants

The circadian clock regulates diverse aspects of plant growth and development and promotes plant fitness. Molecular identification of clock components, primarily in Arabidopsis, has led to recent rapid progress in our understanding of the clock mechanism in higher plants. Using mathematical modeling and experimental approaches, workers in the field have developed a model of the clock that incorporates both transcriptional and posttranscriptional regulation of clock genes. This cell-autonomous clock, or oscillator, generates rhythmic outputs that can be monitored at the cellular and whole-organism level. The clock not only confers daily rhythms in growth and metabolism, but also interacts with signaling pathways involved in plant responses to the environment. Future work will lead to a better understanding of how the clock and other signaling networks are integrated to provide plants with an adaptive advantage.

Respiratory Circuits: Function, Mechanisms, Topology, and Pathology

Neuroscientists have long sought to understand how circuits in the nervous system are organized to generate the precise neural outputs that underlie particular behaviors. Recent studies deepened our understanding of the mechanisms responsible for the generation of the rhythmic output for breathing. Here, the author focuses on issues that are pertinent for the respiratory network and considers its organization and how it derives the functional output. The author discusses pacemaker and network mechanisms of rhythm generation, which are now combined into a novel concept of emergent network activity due to coherent excitation of pacemaker groups. He discusses subcellular basis of this hypothesis and possible mechanisms of synchronization within respiratory network. These new findings in respiratory neuroscience are further applied to explain modifications in breathing during hypoxia and possible origins of respiratory disorders that may be acquired during neural development and aging.