J . Joosse. R . M . Buijs and F . J . H . Tilders (Eds.)

Progress in Brain Research, Vol. 92

0 1992 Elsevier Science Publishers B.V.

All rights reserved

289 CHAPTER 25

Neurotransmitter colocalization and circadian rhythms H. Elliott Albersl, Shyh-Yuh Liou’,*, Edward G. Stopa2 and R. Thomas Zoeller3 I

Laboratory of Neuroendocrinology and Behavior, Departments of Biology and Psychology, Georgia State University, Atlanta, GA 30303, U.S.A., Department of Pathology, State University of New York Health Science Center, Syracuse, N Y 13210, U.S.A. and Department of Anatomy, University of Missouri Medical School, Columbia, MO 65212, U.S.A.

Introduction The timing of behavior and physiological processes is tightly linked to the 24-h day-night cycle. It is thought that this daily rhythmicity evolved as a way of keeping an individual’s behavior and physiology advantageously timed with respect to the external environment. Anticipation and preparation for the cycles which occur in the physical and social environment can provide a powerful adaptive benefit. Circadian clocks are responsible for much of the daily rhythmicity observed in organisms. Two fundamental properties of these clocks keep an individual’s behavior appropriately timed with the environment (Fig. 1). First, circadian clocks are genetically programmed with a cycle length that is slightly longer or shorter than 24 h. This property can be observed in organisms housed in the laboratory when environmental time cues such as the day - night cycle have been eliminated. Under these conditions, a circadian clock is said to be “freerunning” because it is expressing its own intrinsic non-24-h cycle. The length of the circadian cycle varies among species, but does not normally deviate Correspondence: H.E. Albers, Laboratory of Neuroendocrinology and Behavior, Departments of Biology and Psychology, Georgia State University, Atlanta, GA 30303, U.S.A. * Present address: UpJohn Pharmaceuticals Limited, Tsukuba Research Laboratories, 23 Wadai, Tsukuba-shi, 300-42, Japan.

from 24 h by more than a few hours. For example, humans housed in ‘‘free-running” conditions exhibit a circadian cycle of approximately 25 h (Wever, 1979); whereas, many rodent species have “free-running” circadian cycles shorter than 24 h (Pittendrigh and Daan, 1976; Aschoff, 1979). The circadian cycle length remains relatively constant within an individual; however a range of circadian periods can be seen among individuals of the same species. The second essential property of circadian clocks is their ability to become synchronized with the 24-h day - night cycle. The process of synchronization is called “entrainment” and is the result of the clock’s ability to be reset by light. The way in which circadian clocks are reset by the 24-h day-night cycle can be illustrated by observing how brief pulses of light reset circadian clocks under “free-running” conditions. If a rodent housed in constant darkness is exposed to a single 15-min pulse of light around the time that its daily activity begins, its circadian clock will be reset in the delay direction. However, if a 15-min pulse of light is provided 6 h later, more towards the end of the animal’s daily phase of activity, its circadian clock will be reset in the advance direction. If a 15-min pulse of light is provided at other times within the circadian cycle the timing of the clock is not altered. This phase-dependent sensitivity of the clock to light is summarized in a phase response curve (Fig. 2). Since light can reset by advancing or delaying, it



Constant Darkness Ot-----l

LD 1 2 1 2 lo] 20




is possible for light to reset circadianclocks to exactly 24 h. A useful analogy is to think of the circadian clock as a clock that runs one hour too fast (or slow) each day (i.e. with a 25- or 23-h period). If the clock is reset in the advance (or delay) direction each day it can be forced to adopt the 24-h period of the day - night cycle. In this same fashion circadian clocks are reset daily to 24 h and thereby serve as relatively accurate indicators of “time-of-day” .


Fig. 1. Illustration of the two most important functional properties of a circadian clock, its ability to generate circadian rhythms and its ability to become synchronized with the 24-h light - dark cycle. In this example the locomotor activity rhythm of a nocturnal rodent is represented. The 24-h day is divided into an active phase indicated by solid black lines and a rest phase that occurs during the remainder of each day (white). Days are indicated on the vertical axis. During the first 10 days of this record the rodent is housed in constant darkness. Under this condition, the circadian clock expresses its own intrinsic “free-running” cycle because there are no environmental time cues such as a light - dark cycle. In this example the “free-running” circadian cycle is slightly longer than 24 h. On day 10 a light - dark cycle consisting of 12 h light and 12 h darkness (LD 12:12) is imposed. The circadian clock is synchronized or “entrained” to 24 h by the LD cycle. Locomotor activity occurs during the 12 h dark phase (indicated by bracket) in this nocturnally-active rodent.


I Active Phase 12 Time Within Circadian Cycle (hrs)

Rest Phase



Fig. 2. A phase- response curve illustrating the ability of 15-min pulses of light to reset the timing of circadian rhythms in a nocturnal rodent housed in continuous darkness. The circadian cycle is represented on the horizontal axis and divided into rest (i.e. 0- 12) and activity (i.e. 12 - 24) phases. The resetting of effects of light on the circadian clock are indicated on the vertical axis. Light provided around the beginning of the daily active phase resets the clock in the delay direction. Light provided approximately 6 h after the onset of the active phase resets the clock in the advance direction. Light provided at other times in the circadian cycle has little or no effect on the clock.

Localization of a circadian clock in the mammalian brain The search for acircadian clock in the brain of mammals was begun in earnest by Curt Richter (1965). After an exhaustive series of experiments, Richter identified an area in the ventral hypothalamus as a site necessary for the persistence of several circadian rhythms in rats. It was not until 1972, however, that the anatomical location of this clock was more precisely defined. Following the conclusive identification of a direct projection from the retina to the suprachiasmatic nucleus (SCN) of the hypothalamus (Hendrickson, et al., 1972; Moore and Lenn, 1972), destruction of the SCN was found to eliminate the circadian rhythms of locomotor activity and adrenal corticosterone content (Moore and Eichler, 1972; Stephan and Zucker, 1972). Subsequent studies have found destruction of the SCN to eliminate a wide range of circadian rhythms in a variety of mammalian species (Minors and Waterhouse, 1986; Rosenwasser, 1988; Meijer and Rietveld, 1989). Recent evidence from humans indicates that hypothalamic damage that includes the SCN region can also result in the loss of the normal rhythmicity of physiological, behavioral and cognitive variables (Fulton and Bailey, 1929; Gillespie, 1930; Schwartz et al., 1986; Cohen and Albers, 1991) (Fig. 3). The hypothesis that the SCN is a circadian clock has been examined using a variety of experimental approaches (for reviews see Minors and Waterhouse, 1986; Rosenwasser, 1988; Meijer and Rietveld, 1989). Some of the most compelling early evidence that the SCN is capable of generating circa-


Anatomy of the suprachiasmatic nucleus


i i 5 45 6 i S 9 ioiiii i 2 3 4 5 6 i 6




0Awake Sleep

Fig. 3. Unscheduled sleep - wake pattern of a patient with rostra1 hypothalamic damage which includes the suprachiasmatic nucleus. Sleep-wake ratings were recorded every 15 min throughout a one week period by the nursing staff. Days are indicated on the vertical axis and time of day is indicated on the horizontal axis. (From Cohen and Albers, 1991).

dian rhythms came from demonstrations that the SCN exhibited circadian rhythmicity in glucose utilization (Schwartz et al., 1980) and electrical activity (Groos and Hendriks, 1979; Inouye and Kawamura, 1979). For example, the spontaneous discharge of SCN single units exhibit a circadian pattern when recorded in vitro from a small hypothalamic explant. This circadian pattern of single-unit discharge occurs in SCN neurons obtained from rodents maintained in 24-h light-dark cycles and persists in rodents housed in continuous illumination for several months (Fig. 4). More recent evidence in support of the hypothesis that the SCN functions as a circadian clock has been provided by transplantation studies. Circadian rhythmicity can be restored in SCN-lesioned rodents by the transplantation of the SCN, but not other neural tissues (Drucker-Colin et al., 1984; Sawaki et al., 1984; Lehman et al., 1987; DeCoursey and Buggy, 1989; Earnest et al., 1989; Boer and Griffioen, 1990). In fact, some of the characteristics of the circadian clock of the donor can be observed in the host following SCN transplantation. Transplantation of the SCN obtained from mutant hamsters with unusually short circadian rhythms result in the development of short circadian periods in the host (Ralph et al., 1990).

Based on a variety of anatomical criteria the SCN has been subdivided into two predominant subpopulations of neurons, the dorsomedial and ventrolateral (van den Pol, 1980). Neurons within the dorsomedial region are smaller and more tightly packed than those in the ventrolateral area. A large percentage of the neurons within the SCN form local circuits. These interneurons provide the potential for much communication between and within the dorsomedial and ventrolateral subdivisions. It would appear that individual neurons within the SCN can be classified according to their axonal projections: (1) neurons that project only within the SCN, (2) neurons that project only outside the SCN, (3) neurons that project both within the SCN and to sites outside the SCN. Neurons within the dorsomedial and ventrolateral subdivisions produce different neurotransmit-





Fig. 4. Single-unit neuronal activity of SCN neurons recorded extracellularly using the hypothalamic slice preparation. The spontaneous discharge (impulses/s) is indicated on the vertical axis and time within the circadian cycle is indicated on the horizontal axis. A. Firing rates from SCN neurons recorded in a slice obtained from a hamster previously entrained to a light-dark cycle of 14 h light and 10 h darkness. The small arrow indicates that electrical activity peaked at approximately 4.5 h before the onset of the dark phase. B. Firing rates from SCN neurons recorded in a slice obtained from a hamster previously housed in continuous light (LL) for 146 days. The firing rates are plotted as a function of the timing of the circadian locomotor rhythm (CT 1290 = the onset of the active phase). The small arrow indicates that the electrical activity peaked at approximately 9 h before the beginning of the active phase. The dark arrow indicates the time of slice preparation.


ters. Within the dorsomedial division argininevasopressin (AVP) (Vandesande et al., 1975; Card and Moore, 1984; van den Pol, 1986) and somatostatin immunoreactivity (IR) (Dierickx and Vandesande, 1979; Card and Moore, 1984; van den Pol andTsujimoto, 1985) andmRNA(Cardet al., 1988) are found in heavy concentrations. In the ventrolateral subdivision, vasoactive intestinal peptide (VIP) (Loren et al., 1979; Card et al., 1981), peptide histidine isoleucine-27 (PHI) (Stopaet al., 1988)and gastrin releasing peptide (GRP) (van den Pol and Tsujimoto, 1985; Zoeller et al., 1989; Mikkelsen et al., 1991) have been localized (Fig. 5 ) . In addition to neuropeptides, the SCN also contains large numbers

of GABAergic neurons (Card and Moore, 1984; van den Pol, 1986). In fact, based on in situ hybridization studies of glutamic acid decarboxylase (GAD; the synthetic enzyme for GABA) mRNA it has been suggested that GABA may be found within a majority of SCN neurons (Okamura et al., 1989). GABA is the only neurotransmitter so far identified within the SCN that is equally distributed throughout the dorsomedial and ventrolateral subdivisions. A number of other substances have also been localized within cell bodies of the SCN; however, these substances appear to exist in lower concentrations (Moore and Card, 1985; van den Pol and Tsujimoto, 1985).

Fig. 5. Photomicrographs demonstrating the presence of vasoactive intestinal peptide (VIP) and peptide histidine isoleucine (PHI) immunoreactivity within the human and rat suprachiasmatic nuclei (SCN), respectively. A. The rostra1 human SCN is readily defined by Nissl staining. B. VIP immunoreactivity is evident within neurons of the ventral portion of the human SCN (arrows). C. The rat SCN is characteristically ovoid shaped as shown in this Nissl preparation. D. PHI imrnunoreactivity within neurons of the ventral rat SCN (arrows).


Determination of how the various subpopulations of interneurons interact within the SCN awaits a more thorough understanding of the synaptic relationships among SCN neurons (van den Pol, 1980). Existing double labelling studies have shown that GABAergic fibers synapse on SCN neurons that produce VIP, GRP and GABA (van den Pol and Gorcs, 1986; Caste1 et al., 1990; Francois-Bellan et al., 1990). In addition, there is evidence that GRPIR terminals synapse on GRP-IR neurons, that AVP-IR terminals synapse on AVP-IR neurons (van den Pol and Gorcs, 1986), and that somatostatin-IR terminals synapse on VIP-IR neurons (Maegawa et al., 1987). The three major afferent projections to the SCN terminate predominately within the ventrolateral SCN. Two of these pathways are well defined photic inputs to the SCN. The retinal hypothalamic tract (RHT) is a direct projection from retinal ganglion cells to the SCN, and the geniculohypothalamic tract (GHT) is a secondary projection from the intergeniculate leaflet of the thalamus (IGL) to the SCN (Hendrickson et al., 1972; Moore and Lenn, 1979; Swansonet al., 1974; Ribak and Peters, 1975; Pickard, 1982). The neurotransmitter contained in the RHT has not been clearly defined; however, there is evidence that the neurotransmitters in this pathway may include glutamate or N-acetylaspartylglutamate (NAAG) (Liou et al., 1986; Shibata et al., 1986; Meijer et al., 1988; Moffett et al., 1990). Several lines of evidence suggest that neuropeptide Y (NPY) functions as the neurotransmitter within the GHT (Card et al., 1983; Albers et al., 1984; Albers and Ferris, 1984; Moore et al., 1984; Harrington et al., 1985). In addition, a major serotonergic projection from the midbrain raphe terminates within the ventrolateral SCN (Fuxe, 1965; Aghajanian et al., 1969; Saavedra et al., 1974). In summary, the SCN receives the majority of its afferent input through the ventrolateral subdivision. Although it is clear that both the RHT and GHT communicate photic information to the SCN (for a review see Meijer, 1991), the type of lighting information that is communicated via these projections is not fully understood. Light primarily activates

photically i.;gen cells within the SCN, although some cells are light-suppressed. Electrical stimulation of the GHT (Rusak et al., 1989) and microinjection of NPY into the SCN (Albers and Ferris, 1984) produce phase shifts in circadian rhythms that mimic the phase shifts produced by exposing animals to pulses of darkness (Boulos and Rusak, 1982; Ellis et al., 1982) or transitions from light to darkness (Albers, 1986). In contrast, electrical stimulation of the RHT produces a pattern of phase shifts that mimics the phase shifts produced by pulses of light (Shibata and Moore, 1989). There is also some evidence that the serotonergic projection from the raphe to the SCN may be involved in the SCN responsiveness to photic stimulation (for a review see Albers et al., 1991a). Thus, it would seem that the three major projections to the SCN may communicate different types of information about environmental lighting to the SCN. If so, it will be important to understand how afferent photic information necessary for the synchronization of circadian rhythms is integrated within the SCN. Terminals of all three major pathways to the SCN appear to terminate directly on SCN neurons that produce VIP. Following optic enucleation, degenerating axons were found to synapse on SCN neurons exhibiting VIP-IR thus suggesting that the RHT synapses directly on VIP-IR neurons (Ibata et al., 1989). Double labelling studies have found 3H serotonin uptake sites (Kiss et al., 1984; Bosler and Beaudet, 1985) and NPY-IR terminals (Hisano et al., 1988) in apposition with VIP-IR dendrites and cell bodies within the SCN. Terminals of the RHT, GHT and serotonergic projection also appear to synapse on other SCN neurons that are not immunopositive for VIP and are yet to be chemically defined. There is also evidence that individual SCN neurons can receive more than one type of afferent input. Serotonergic and NPY, as well as serotonergic and GABAergic terminals have been found to converge on the same dendrites (Guy et al., 1987; Bosler, 1989). The efferent projections of the SCN have been investigated using several different techniques (for a review see Watts, 1991). It has been estimated that


75% of SCN efferents are contained in a pathway that terminates in a region just ventral to the paraventricular nucleus, called the sub-paraventricular zone (sPVNz) (Watts et al., 1987). Combined immunocytochemistry and retrograde tracing have shown that AVP, VIP and a few neurotensin-IR fibers project from the SCN to the sPVNz (Watts and Swanson, 1987). Immunocytochemical evidence indicates that GRP-IR neurons also send heavy projections to the sPVNz, as well as other CNS sites (Mikkelsen et al., 1991).

Colocalization within the suprachiasmatic nucleus It is now well established that multiple neurotransmitters can be colocalized within the same neuron (for a reveiw see Hokfelt et al., 1986). Several different forms of neurotransmitter colocalization have been identified. Classical neurotransmitters can coexist with other classical neurotransmitters, or with peptides; alternatively, peptides can coexist with other peptides. Variations also exist in the molecular mechanisms responsible for the production of colocalized neurotransmitters (Sofroniew et al., 1984). For example, colocalization can result when multiple peptides are processed from a common precursor molecule, or when different neurotransmitter genes are expressed within the same neuron. Within the SCN, several different forms of neurotransmitter colocalization are observed. VIP and PHI are colocalized because they are derived from a common polypeptide precursor (Nishizawa et al., 1985). VIP and PHI are thought to be cleaved from the precursor by proteolytic processing. Both VIPand PHI-IR appear within the SCN early in development around day 18 - 20 of gestation (Ishikawa and Frohman, 1987; Laemle, 1988). Existing evidence suggests that the post-translational processes necessary for the synthesis of VIP and PHI are regulated in a similar manner since VIP- and PHI-IR occur in approximately a 1:l ratio within the SCN under several different conditions (Albers et al., 1987). However, there is data to suggest that different subpopulations of VIP-producing neurons

could release different forms of PHI (Cauvin et al., 1991). Recent evidence indicates that SCN neurons that produce VIP/PHI can also colocalize other neurotransmitters. Immunocytochemical studies have identified GRP within VIP/PHI producing neurons of the SCN (Okamura et al., 1986). It seems likely that VIP/PHI and GRP can all be synthesized within at least some SCN neurons since the mRNAs encoding VIP/PHI and GRP can be colocalized within SCN neurons (Albers et al., 1991b). More recently double labelling studies have identified both GAD and VIP within axonal varicosities of the SCN indicating the probable coexistence of VIP/PHI and GABA within SCN neurons (Francois-Bellan et al., 1990). Interestingly, a novel 712 amino acid protein designated VGF (van den Pol et al., 1989) can be colocalized in VIP- and in AVP-IR neurons within the SCN. The possible functions of VGF in circadian control remain unknown. In summary, VIP-producing neurons within the SCN appear to be a heterogenous population of cells. SCN neurons that produce VIP can be classified based on the number and type of neurotransmitters with which VIP coexists (Table I). The TABLE I Potential complexity of vasoactive intestinal peptide (VIP) producing neurons in the SCN. SCN neurons that produce VIP could fall into any one of 72 possible combinations of afferent terminals, colocalized neurotransmitters and efferent projections. The potential complexity of these neurons will increase when the synaptic contacts of their efferent projections are defined. Abbreviations: RHT, retinohypothalamic tract; GHT, geniculohypothalamic tract; RAPHE, serotonergic projection from the raphe to the SCN. Afferents





Within SCN Extra SCN Both


potential number of different classes of VIP neurons increases substantially if one also considers the different patterns of afferent input and efferent projections that can be displayed by individual neurons. Demonstration of the percentage of SCN neurons that fall into all these possible classes of VIP neurons will be important, although extremely difficult given the number of antigens that need to be simultaneously examined. Identification of the potential physiological role of the various classes of VIP-containing SCN neurons should be useful in focusing the anatomical investigation of these neurons.







--% -?

Function of colocalized SCN neurotransmitters The release of multiple neurotransmitters in what has been termed a “cocktail” (Swanson, 1983) or “bouquet” (Greenberg and Price, this volume) has the potential to vastly increase the signalling capabilities of a neuron. Little is known about how the chemical composition of these cocktails is regulated, or how different cocktails of neurotransmitters may influence their targets. The SCN provides a useful neuronal system in which to study these mechanisms because its intrinsic anatomy has been studied intensively, it has well defined inputs (i.e. RHT, GHT and serotonergic projection from the raphe) and easily quantifiable outputs (e.g. circadian rhythms). We have investigated the response of the SCN to VIP, PHI and GRP, as well as the factors that regulate the availability of these peptides for release from SCN neurons.

Interactive effects of VIP, PHI and GRP within the SCN T o investigate the potential functional consequences of the corelease of VIP, PHI and GRP from local circuits within the SCN we examined whether the microinjection of a cocktail containing equimolar concentrations of VIP, PHI and GRP (VIP/ PHI/GRP) into the SCN could influence circadian timekeeping (Albers et al., 1991b). The microinjection of VIP/PHI/GRP delayed the phase of free-

Fig. 6 . Effect of administration of various combinations of VIP, PHI and GRP on the free-running locomotor rhythm of hamsters following microinjection into the suprachiasmatic region. Coadministration of VIP, PHI and GRP produced large phase delays when given around the time of the onset of the active phase, but not at other times in the circadian cycle. All microinjections contained the same final concentration of total peptide. Closed circles indicate time of injection. (From Albers et al., 1991bl.

running hamster circadian rhythms when administered at the beginning of the daily active phase, but not at other times within the circadian cycle (Figs. 6 and 7 ) . Comparison of the effects of VIP/PHI/ GRP on circadian timing with the effects of light suggests that coadministration of these peptides mimics the phase delaying, but not the phase advancing effects of light (compare Figs. 2 and 7). The phase-delaying effects of these peptides on circadian rhythms appear to be the result of their action within the SCN region since their injection into the cerebroventricular system does not alter circadian timing. Since coadministration of VIP, PHI and GRP mimics the phase-delaying effects of light, one or more of these peptides may play a role in the synchronization of circadian rhythms with the lightdark cycle. Although administration of a cocktail containing VIP, PHI and GRP could produce phase delays, it




Circadian time

Fig. 7. A phase-response curve illustrating the ability of microinjections (n = 39) of equimolar concentrations of VIP/ PHI/GRP into the suprachiasmatic region to reset the timing of circadian rhythms. Solid circles represent mean phase shifts (k S.E.M.), in hours, during 3-h intervals throughout the circadian cycle (circadian time 12 refers to the beginning of the active phase). VIP/PHI/GRP produced phase delays around the time of activity onset, but had little effect at other times in the circadian cycle. Comparison of these data with the phase shifting effects of light (Fig. 2) indicate that VIP/PHI/GRP microinjections into the SCN mimic the phase delaying, but not the phase advancing effects of light. (From Albers et al., 1991b).

remained necessary to determine whether the combined administration of these peptides was necessary for the production of these circadian effects. As a result the phase delaying effects of administration of VIP, PHI and GRP individually, or coadministration of VIP/PHI, VIP/GRP and PHI/GRP were compared with the effects of VIP/PHI/GRP. The final concentration of total peptide in each injection was the same. As can be seen in Fig. 8 (top) only the combined administration of VIP, P H I and GRP produced maximal phase delays of approximately 1.5 h. The administration of each peptide alone, or in combination with one other produced maximal phase delays that were at least 50% smaller than coadministration of all three peptides in combination. The interactive effects of VIP, PHI and GRP on SCN neurons could also be observed at the cellular level using the hypothalamic slice preparation (Albejs et al., 1991b). Combined administration of VIP/PHI/GRP produced a significant increase in

the spontaneous discharge rate of SCN single units recorded extracellularly (Fig. 9). In contrast, administration of each peptide alone, or in combination with one of the other peptides, produced a smaller increase in firing rate (Fig. 8, bottom). In summary, studies at both the behavioral and cellular level indicate that VIP, PHI and GRP interact to produce maximal effects on circadian rhythms within the SCN. These data suggest that the corelease of a cocktail containing VIP, PHI and GRP would produce significantly different effects on circadian rhythms, than the release of each peptide alone, or each peptide coreleased with one other. In addition, the way in which VIP, PHI and GRP interact appears to illustrate a novel synaptic mechanism in which each coreleased neurotransmitter contributes equally to the production of the functional response. Factors regulating VIP, PHI and GRP within SCN neurons

Several lines of evidence now suggest that environmental lighting conditions significantly influence SCN neurons that produce VIP and PHI. VIP- and PHI-IR are significantly lower in the SCN of rats housed in continuous light as compared to continuous darkness (Albers et al., 1987). Light does not have similar effects on VIP- and PHI-IR in cortex, or on substance P- or neurotensin-IR within the SCN. Further support for the possibility that photic afferents influence VIP-IR within the SCN has come from studies where enucleation was found to increase SCN levels of VIP-IR and mRNA (Okamura et al., 1987; Holtzman et al., 1989). The serotonergic projection from the raphe may also influence SCN levels of VIP. Depletion of serotonin reduces VIP-, but not AVP- or NPY-IR within the SCN (Kawakami et al., 1985; Guy et al., 1987). There is also a 24-h rhythm in the levels of the mRNA encoding VIP and PHI within the SCN. Studies using a variety of hybridization techniques including quantitative in situ, solution and dot-blot hybridization have demonstrated that SCN levels of VIP/PHI mRNA are lower during the day than at



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Neurotransmitter colocalization and circadian rhythms.

J . Joosse. R . M . Buijs and F . J . H . Tilders (Eds.) Progress in Brain Research, Vol. 92 0 1992 Elsevier Science Publishers B.V. All rights res...
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