Location of the reproductive timer in the male cricket Gryllus bimaculatus DeGeer as revealed by local cooling of the central nervous system

The location of the reproductive timer for the post-copulatory, time-fixed, sexually refractory stage was investigated in the male cricket Gryllus bimaculatus. This stage was defined as the interval between spermatophore protrusion and recommencement of copulation or a calling song. To inactivate the central nervous system locally and reversibly, different body regions were cooled to 10°C for 20-30 min after spermatophore protrusion. A behavioural test then measured the duration of the refractory stage after males recovered from cooling. Males with the head, thorax and anterior abdomen cooled did not show a lengthening of that stage. In contrast, males with the entire abdomen or even the posterior abdominal segments containing only the 6th and terminal (7th-11th) abdominal ganglia showed a lengthening of the refractory stage up to, but not exceeding, the cooling duration. When 20-min cooling was interposed twice after spermatophore protrusion, the refractory stage was lengthened by about 40 min, indicating that interposed cooling did not reset the timer. These results are in agreement with our previous hypothesis that the reproductive timer for the refractory stage in the male cricket is located in the posterior abdominal ganglia, possibly within the terminal abdominal ganglion.     

Location of the reproductive timer in the male cricket Gryllus bimaculatus DeGeer as revealed by local cooling of the central nervous system.

The location of the reproductive timer for the post-copulatory, time-fixed, sexually refractory stage was investigated in the male cricket Gryllus bimaculatus. This stage was defined as the interval between spermatophore protrusion and recommencement of copulation or a calling song. To inactivate the central nervous system locally and reversibly, different body regions were cooled to 10 degrees C for 20-30 min after spermatophore protrusion. A behavioural test then measured the duration of the refractory stage after males recovered from cooling. Males with the head, thorax and anterior abdomen cooled did not show a lengthening of that stage. In contrast, males with the entire abdomen or even the posterior abdominal segments containing only the 6th and terminal (7th-11th) abdominal ganglia showed a lengthening of the refractory stage up to, but not exceeding, the cooling duration. When 20-min cooling was interposed twice after spermatophore protrusion, the refractory stage was lengthened by about 40 min, indicating that interposed cooling did not reset the timer. These results are in agreement with our previous hypothesis that the reproductive timer for the refractory stage in the male cricket is located in the posterior abdominal ganglia, possibly within the terminal abdominal ganglion.     

Auto-spermatophore extrusion reveals that the reproductive timer functions in the separated terminal abdominal ganglion in the male cricket

Auto-spermatophore extrusion is a kind of spermatophore extrusion without genital coupling in the male cricket. It rarely occurred in intact males paired with a female, while it frequently occurred in all the males with the connectives cut under restraint and dissection. The time interval (SPaSE) between spermatophore preparation and auto-spermatophore extrusion was found to be comparable to that (RS2) of the time-fixed sexually refractory stage measured by the calling song. According to extracellular spike recording, the dorsal pouch motoneuron (mDP), which singly innervates the dorsal pouch muscles and is responsible for spermatophore extrusion, showed a burst discharge in association with auto-spermatophore extrusion with an interval similar to RS2 in males with the connectives transected between the 6th abdominal ganglion and the terminal abdominal ganglion (TAG) after spermatophore preparation. These results strengthened our previous conclusion that the reproductive timer for RS2 is located in the TAG, and demonstrated that it functions normally even in the TAG separated from the rest of the central nervous system.     

Thee effect of 5-HTP on the reproductive timer in the male cricket

The post-copulatory sexually refractory stage in the male cricket Gryllus bimaculatus consists of the two substages: the first refractory stage (RS1, time-variable) between copulation and spermatophore preparation, and the second refractory stage (RS2, time-constant) between spermatophore preparation and the recommencement of courtship. To understand the mechanism of the timer for RS2, subcuticular or intraganglionic injection of biogenic amines (10(-2) mol l(-1)) was performed immediately after spermatophore preparation. RS2 was shortened by octopamine, 5-HT, 5-HTP and NA-5-HT. Among these, 5-HTP was most potent. It shortened RS2 to maximally about 38% of the control. The shortening effect continued for 4.5 h after subcuticular injection even when the hemolymph was washed out with saline at 1 hour after injection. Simultaneous injection of 5-HTP with the inhibitor (NSD-1015) of 5-HT synthesis enzyme nullified the effect of 5-HTP, indicating that the shortening effect was caused by 5-HT synthesized from extrinsic 5-HTP. Injection of the inhibitor (CHX) of protein synthesis had no effect of on RS2. These results suggest that the reproductive timer in the TAG may be controlled by 5-HT or a second messenger mediated by 5-HT.     

Different effects of the biogenic amines dopamine,serotonin and octopamine on the thoracic and abdominal portions of the escape circuit in the cockroach

1. The escape behavior of the cockroach, Periplaneta americana, is known to be modulated under various behavioral conditions (Camhi and Volman 1978; Camhi and Nolen 1981; Camhi 1988). Some of these modulatory effects occur in the last abdominal ganglion (Daley and Delcomyn 1981a, b; Libersat et al. 1989) and others in the thoracic ganglia (Camhi 1988). Neuromodulator substances are known to underlie behavioral modulation in various animals. Therefore, we have sought to determine whether topical application of putative neuromodulators of the escape circuit enhance or depress this circuit, and whether these effects differ in the last abdominal vs. the thoracic ganglia. 2. Topical application of the biogenic amines serotonin and dopamine to the metathoracic ganglion modulates the escape circuitry within this ganglion; serotonin decreases and dopamine enhances the response of leg motoneurons to activation of interneurons in the abdominal nerve cord by electrical or wind stimulation. 3. The neuropil of the thoracic ganglia contains many catecholamine-histofluorescent processes bearing varicosities, providing a possible anatomical substrate for dopamine release sites. 4. Topical application of octopamine to the terminal abdominal ganglion enhances the response of abdominal interneurons to wind stimulation of the cerci. In contrast, serotonin and dopamine have no effect at this site. 5. It is proposed that release of these biogenic amines may contribute to the known modulation of the cockroach escape response.     

Putative neurohemal areas in the peripheral nervous system of an insect,Gryllus bimaculatus,revealed by immunocytochemistry

The morphology and position of putative neurohemal areas in the peripheral nervous system (ventral nerve cord and retrocerebral complex) of the cricket Gryllus bimaculatus are described. By using antisera to the amines dopamine, histamine, octopamine, and serotonin, and the neuropeptides crustacean cardioactive peptide, FMRFamide, leucokinin 1, and proctolin, an extensive system of varicose fibers has been detected throughout the nerves of all neuromeres, except for nerve 2 of the prothoracic ganglion. Immunoreactive varicose fibers occur mainly in a superficial position at the neurilemma, indicating neurosecretory storage and release of neuroactive compounds. The varicose fibers are projections from central or peripheral neurons that may extend over more than one segment. The peripheral fiber varicosities show segment-specific arrangements for each of the substances investigated. Immunoreactivity to histamine and octopamine is mainly found in the nerves of abdominal segments, whereas serotonin immunoreactivity is concentrated in subesophageal and terminal ganglion nerves. Immunoreactivity to FMRFamide and crustacean cardioactive peptide is widespread throughout all segments. Structures immunoreactive to leucokinin 1 are present in abdominal nerves, and proctolin immunostaining is found in the terminal ganglion and thoracic nerves. Codistribution of peripheral varicose fiber plexuses is regularly seen for amines and peptides, whereas the colocalization of substances in neurons has not been detected for any of the neuroactive compounds investigated. The varicose fiber system is regarded as complementary to the classical neurohemal organs.     

Presynaptic effects of octopamine, serotonin, and cocktails of the two modulators on neuromuscular transmission in crustaceans

The effect of the biogenic amines octopamine and serotonin, and of both amines combined (cocktails) on transmitter release at neuromuscular junctions of two crustaceans was studied. octopamine (10(-8) mol l(-1) to 10(-6) mol l(-1)) either enhanced or decreased evoked transmitter release through presynaptic effects. The results were identical for the slow and the fast excitor in the closer muscle of the crab, and for the excitor in the opener muscle of the crayfish. Application of serotonin always resulted in a strong increase of release. However, this potentiating effect of serotonin was reduced in strength by subsequent application of cocktails consisting of serotonin and octopamine. In all experiments, a cocktail of serotonin and octopamine was less effective than serotonin alone. The decrease in the mean quantal content m by octopamine was due to a reduction of the probability of release p. Since both amines are synthesized in the central nervous system and are released from neurohaemal organs into the haemolymph bathing the neuromuscular junctions, the results suggest that the two amines, when present together, modulate transmitter release in an antagonistic way, and that the level of the two determines synaptic efficacy.     

Aminergic cellular organization in the gills of Aplysia species

The constituent elements of the gills of Aplysia kurodai and A. juliana were examined for the presence of biogenic amines using histochemical, immunocytochemical, and HPLC techniques. Aminergic elements were revealed by glyoxylic acid-induced fluorescence in the branchial nerve, branchial ganglion, branchial vessels, and pinnules in both species. Three types of fluorescent cells were found in the neural plexus of the gill in each species. Two of them might be sensory neurons. Although HPLC analysis showed the presence of serotonin and dopamine in all gill structures including fluorescent neural elements, there were regional differences in concentrations of the monoamines. It was noted in the pinnules that there was a much higher concentration of dopamine than serotonin. Serotonin immunocytochemistry revealed neural processes which were immunoreactive to antiserotonin antibody, but serotonin immunoreactivity could not be found in a population of branchioganglionic neuron (BGN) somata. Serotonergic elements in the ganglion may be processes of the central ganglion, while dopaminergic elements may be processes of neurons in the neural plexus, located beyond the branchial ganglion. BGNs were activated by bath-applied dopamine and serotonin. These results suggest that dopaminergic sensory inputs from the neural plexus and serotonergic descending inputs from the abdominal ganglion may be among the inputs received by BGNs. It was found that serotonin depressed excitatory junctional potentials in muscle cells of the efferent branchial vessel, which were induced by an identified neuron of the abdominal ganglion. The aminergic cellular organization of the gill may involve serotonergic presynaptic-inhibitory fibers arising from the abdominal ganglion.     

Cellular and molecular mechanisms controlling melatonin release by mammalian pineal glands

1. The pineal gland is regulated primarily by photoperiodic information attaining the organ through a multisynaptic pathway initiated in the retina and the retinohypothalamic tract. 2. Norepinephrine (NE) released from superior cervical ganglion (SCG) neurons that provide sympathetic innervation to the pineal acts through alpha1- and beta 1- adrenoceptors to stimulate melatonin synthesis and release. 3. The increase in cyclic AMP mediated by beta 1-adrenergic activation is potentiated by the increase in Ca2+ flux, inositol phospholipid turnover, and prostaglandin and leukotriene synthesis produced by alpha 1-adrenergic activation. 4. Central pinealopetal connections may also participate in pineal control mechanisms; transmitters and modulators in these pathways include several neuropeptides, amino acids such as gamma-aminobutyric acid (GABA) and glutamate, and biogenic amines such as serotonin, acetylcholine, and dopamine. 5. Secondary regulatory signals for pineal secretory activity are several hormones that act on receptors sites on pineal cells or at any stage of the neuronal pinealopetal pathway.     

Internalization of seasonal time

Orientation of physiology and behavior in time is a major adaptation common to many organisms and represents a significant challenge. In the face of predictable seasonal changes in climatic factors, and as a result of natural selection, many mammals now restrict their reproductive efforts to the fraction of the year when conditions of temperature, food, and water are most favorable for successful weaning of offspring. Changes in day length figure prominently in the synchronization of mammalian seasonal reproductive cycles. In summer breeders (e.g., several hamster, vole, and mouse species), decreasing summer day lengths induce reproductive involution. During this interval, neuroendocrine mechanisms analogous to a simple reference memory permit discrimination of stimulatory from inhibitory photoperiods. The non-reproductive phenotype is sustained for several months thereafter by the inhibitory short days of late summer, autumn, and early winter. Mid-winter reactivation of the reproductive system is triggered after 20-25 weeks of exposure to decreasing or short days by an interval timer that renders the reproductive neuroendocrine system refractory to short days. Recent work that has explored formal and physiological properties of this photorefractoriness interval timer has identified sex differences, neural substrates, and changes in hypothalamic gene expression that may participate in the measurement of seasonal time.     

Neurohormonal control of the mating interval in the male cricket,Gryllus bimaculatus DeGeer

Summary Between two mating acts of the male cricket (Gryllus bimaculatus), spermatophore protrusion (SP) and courtship stridulation (CS), there is a fixed time interval. This interval lasts about 1 h. During the period from SP to CS, the male cricket does not stridulate nor make any type of mating sound (post-spermatophore protrusion silence: PSPS) and tolerates external sensory stimuli. We examined the effects of injections of hemolymph and ganglia extracts on the interval. Extracts obtained from crickets which had just started CS (CS crickets) and those which had finished SP (SP crickets) were effective. The extracts were fractionated by ul trafiltration. Fractions with a molecular weight of less than 1 kdalton affected the length of the PSPS. The fractions from both the hemolymph and the mesothoracic ganglion of CS crickets shortened the PSPS. On the other hand, the fractions from the hemolymph and the brain of SP crickets lengthened the PSPS. We estimated, by gel filtration, the molecular weight of the effective fractions from the mesothoracic ganglion and the brain to be 100–200 daltons. We also examined the effects of biogenic amines on the PSPS. Octopamine shortened the PSPS, whereas serotonin lengthened it. The results in dicate that at least two neurohormones from the brain and the mesothoracic ganglion reciprocally control the elicitation of CS and provide an appropriate interval in the mating sequence of the male cricket. Octopamine and serotonin are possible candidates for these neurohormones.Abbreviations CP copulation - CS courtship stridulation - SP spermatophore protrusion - PSPS post-spermatophore protrusion silence     

Effects of octopamine, dopamine, and serotonin on production of flight motor output by thoracic ganglia of Manduca sexta

Effects of biogenic amines on a centrally generated motor pattern in Manduca sexta were examined by pressure injecting nanomole to micromole amounts of octopamine, dopamine or serotonin into thoracic ganglia. Motor output was recorded extracellularly from a pair of antagonistic flight muscles and their motor neurons. The monoamines were found to alter production of a motor pattern that produces rhythmic wing flapping (10 Hz) and exhibits phase relationships similar to those in the flight pattern of intact moths. In mesothoracic ganglia with sensory nerves intact, octopamine (4 X 10(-9) mol) injected into lateral regions evoked regular firing of a single motor neuron, whereas a higher dose (4 X 10(-8) mol) often elicited the flight motor pattern. In the absence of sensory input, these doses of octopamine had little effect. Low doses (10(-10) mol) greatly enhanced motor responses to electrical stimulation of a wing sensory nerve. Dopamine (2 X 10(-10) mol) injected into the medial region of the mesothoracic ganglion elicited the flight motor pattern in the presence or absence of sensory input. Rhythmic output induced by dopamine (5 X 10(-10) mol) was suppressed by injecting serotonin (5 X 10(-10) mol) into the same region. These findings demonstrate that dopamine, octopamine, and serotonin have different effects on motor output in Manduca and suggest that these amines are involved in initiating, maintaining and terminating flight behavior, respectively. Octopamine may elicit flight production by enhancing the efficacy of sensory transmission thereby increasing excitability or arousal. Dopamine may act on interneurons involved in generating the flight motor pattern.     

NADPH-diaphorase histochemistry in the terminal abdominal ganglion of the crayfish

Nitric oxide (NO) has an important modulatory role on the processing of sensory signals in vertebrates and invertebrates. In this investigation we studied the potential sources of NO in the terminal abdominal ganglion of the crayfish, Pacifastacus leniusculus, using NADPH-diaphorase (NADPHd) histochemistry, with NADPHd acting as a marker for NO synthase (NOS). In the terminal ganglion a mean of 27 strongly labelled NADPHd-positive cell bodies were found, and of these 80% [of stained cell bodies] [corrected] occurred in three regions located in antero-lateral, central and posterior parts of the ganglion. Ventral and antero-ventral commissures as well as specific dorsal and ventral areas of the dendritic neuropil showed positive staining. Intense labelling was seen in the ventro-medial tract, and in the connective between the terminal ganglion and the 5th abdominal ganglion. In addition, some motor neurones and neurones with branches in the sensory commissures were NADPHd positive. Our finding that NADPHd-positive cells occur in consistent patterns in the terminal abdominal ganglion implies that NO may have a role in mechanosensory processing in the crayfish.     

Brain control of mating behavior in the male cricket Gryllus bimaculatus DeGeer: brain neurons responsible for inhibition of copulation actions

We localized brain neurons responsible for the inhibition of copulation actions in the male cricket, Gryllus bimaculatus. Males with one connective-cut between the brain and the terminal abdominal ganglion (TAG) were able to perform normal reproductive behaviors such as courtship, copulation and spermatophore protrusion. However, copulation response tests using a female dummy showed that they failed to exhibit a transient inhibitory state after application of noxious stimulation in the mating stage and to exhibit a tonic inhibitory state in the post-copulatory sexual refractory stage. A partial cut of a single connective (hemilateral connective-cut) combined with the copulation response test and an axonal backfilling demonstrated that approximately 40 brain neurons (per one connective), whose somata were located in the posterior region of the protocerebrum and mainly contralaterally to their descending axons running through the dorso-medial part of the connective, were candidates for the inhibition of copulation actions, at least in the mating stage. In addition, we report here for the first time, that copulation actions can be elicited shortly after copulation even in intact males. This unusual behavior was frequently observed in fledglings after the final molt. The results are discussed from the perspective of the operation of the brain inhibition system on the central pattern generator (CPG) for copulation.     

Amine modulation of the neurogenic Limulus heart

(1) The biogenic amines octopamine (OCT), dopamine (DA), epinephrine (E), and norepinephrine (NE) cause dose-dependent increases in both the rate and amplitude of contractions of the isolated Limulus heart-cardiac ganglion. Their relative ability to produce this excitation is OCT greater than DA approximately the same as E greater than NE. (2) The excitatory effects of all these amines are antagonized by the alpha-adrenergic blocker phentolamine and the dopaminergic antagonist haloperidol. The beta-adrenergic antagonist dichloroisoproterenol slightly reduces amine excitation, but is also a partial agonist. The beta-adrenergic antagonist propanolol, the alpha-blocker phenoxybenzamine, and the serotonin antagonist metergoline are ineffective. (3) In addition to their excitatory effects, DA and, to a lesser extent, NE initially reduce contraction rate and amplitude. (4) The transient inhibition is eliminated selectively by metergoline and is unaffected by the other antagonists. (5) The amines all increase the frequency of cardiac ganglion electrical bursting activity, whether ganglia are isolated or attached to cardiac muscle. Dopamine and NE also transiently inhibit the cardiac ganglion. (6) The amines do not alter myocardial resting tension, contractility, or membrane potential. (7) These amines appear to exert their modulatory effects on Limulus heart by altering the properties of the neurons which comprise its cardiac ganglion.     

Baclofen modifies via the release of monoamines the synaptic depression, frequency facilitation, and posttetanic potentiation observed at an identified cholinergic synapse of Aplysia californica

1. The mechanism of action of baclofen was studied at a cholinergic synapse of Aplysia. This synapse, called RC1-R15, can be activated by a minimal stimulation of the right pleurovisceral connective and is recorded in cell R15 of Aplysia californica. Repeated stimulation of synapse RC1-R15 produces depression, followed by frequency facilitation and by posttetanic potentiation (PTP). 2. Perfusion with baclofen (3 x 10(-5) or 5 x 10(-5) M) reduces the size of all excitatory postsynaptic potentials (EPSPs) of synapse RC1-R15 produced by a train of 100 stimuli at 1.5 Hz. It also reduces the synaptic depression and PTP but increases the frequency facilitation. These effects are similar to those produced by gamma-aminobutyric acid (GABA), serotonin, and dopamine on this synapse. 3. When the preparation is perfused with bicuculline or picrotoxin, two antagonists of GABA, the effects of baclofen are not antagonized. However, when baclofen is perfused in the presence of SQ10,631 (an antagonist of serotonin) or butaclamol (an antagonist of dopamine), its effects are partially blocked. 4. To determine if baclofen produces its action by direct interaction with aminergic receptors or by liberating the amines from some nerve endings, a few animals were treated with reserpine to deplete the aminergic pool. Following this treatment no effects were obtained with baclofen suggesting that it acts by liberating dopamine and serotonin or some other amines. 5. In animals treated with reserpine, GABA still produces its normal effects (reduction of EPSP size, synaptic depression, posttetanic potentiation, and increase of facilitation), indicating that baclofen does not act directly on the GABA receptor located on the presynaptic terminal.     

Effect of Acute Fluoxetine Treatment on the Brain Serotonin Synthesis as Measured by the α-Methyl-l-Tryptophan Autoradiographic Method

Abstract: The effect of treatment with acute fluoxetine, a serotonin reuptake inhibitor, on the rate of serotonin synthesis in the rat brain was studied through autoradiography following intravenous administration of α-methyl-l -[³H]tryptophan. The rate of serotonin synthesis in fluoxetine-treated rats was compared with the rate measured in sham-treated rats (saline injection). Results showed a significant increase in the rate of synthesis in the majority of cerebral structures examined. The greatest increase (given as a percentage of rates in control animals) in the rate of serotonin synthesis was observed in the substantia nigra compacta (344%), hippocampus-CA3 (337%), dorsal hippocampus (283%), and caudate-putamen (232%). Fluoxetine had a less significant effect on the rate of synthesis in the pineal body (44%). Data suggest that acute fluoxetine treatment (30 mg/kg, i.p.) enhances the rate of serotonin synthesis in all the structures of rat brain examined in this work.     

Biogenic amines regulate the reproductive function in <Emphasis Type="Italic">Drosophila</Emphasis> as neurohormones

We studied the influence of experimental increase in the octopamine and dopamine content on the level of juvenile hormone degradation, oogenesis, and fertility in wild type Drosophila virilis flies. Feeding of flies on octopamine led to a significantly decreased level of juvenile hormone degradation (increased titer) in young and sexually mature females, rather than in males, markedly decreased the number of vitellogenic (stages 8–10) and mature (stage 14) oocytes), and sharply reduced fertility. Feeding of flies on dopamine decreased the juvenile hormone degradation (increased titer) in young wild type females and increased it (lowered the juvenile hormone titer) in sexually mature females, as well as decreased the fertility of wild type females to a level characteristic for D. virilis line with a mutation doubling the endogenous dopamine level. A possible mechanism of the influence of these amines on the reproductive function in Drosophila as neurohormones is discussed and a conclusion is drawn that the reduced fertility of females at an increased level of amines appears to be related to an increased level of ecdysteroids, which is caused by an increased, as a result of decreased degradation, juvenile hormone titer.     

Octopamine-like immunoreactivity in the dorsal unpaired median (DUM) neurons innervating the accessory gland of the male cockroach Periplaneta americana

The musculature of the mushroom-shaped accessory gland receives innervation from trunks 5C1 of the phallic nerves, which arise from the posterior part of the terminal abdominal ganglion of the male cockroach Periplaneta americana. Anterograde cobalt filling through trunks 5C1 with the subsequent precipitating procedure has shown the fine innervation of the accessory gland. By retrograde cobalt filling through the same trunks, different types of cells have been mapped in the terminal abdominal ganglion. About 25 dorsal unpaired median (DUM) neurons have been identified among them. About 36 octopamine-like immunoreactive DUM neurons with large somata have been characterized in whole-mount preparations of the terminal abdominal ganglion. The combination of the cobalt-filling technique with immunohistochemical mapping of cells suggests an octopaminergic innervation of the musculature of the accessory gland by DUM neurons.