Neurosecretory Process Research Articles

Sympathomimetics regulate quantal acetylcholine release at neuromuscular junctions through various types of adrenoreceptors

The studies of the interaction between the sympathetic and motor nervous systems are extremely relevant due to therapy for many neurodegenerative and cardiovascular disorders involving adrenergic compounds. Evidences indicate close contact between sympathetic varicosities and neuromuscular synapses. This raises questions about the effects of catecholamines on synaptic transmission. The currently available information is contradictory, and the types of adrenoreceptors responsible for modulation of neurotransmitter release have not been identified in mammalian neuromuscular synapses. Our results have shown that the α1A, α1B, α2A, α2B, α2C, and β1 adrenoreceptor subtypes are expressed in mouse diaphragm muscle containing neuromuscular synapses and sympathetic varicosities. Pharmacological stimulation of adrenoreceptors affects both spontaneous and evoked acetylcholine quantal secretion. Agonists of the α1, α2 and β1 adrenoreceptors decrease spontaneous release. Activation of the α2 and β1 adrenoreceptors reduces the number of acetylcholine quanta released in response to a nerve stimulus (quantal content), but an agonist of the β2 receptors increases quantal content. Activation of α2 and β2 adrenoreceptors alters the kinetics of acetylcholine quantal release by desynchronizing the neurosecretory process. Specific blockers of these receptors eliminate the effects of the specific agonists. The action of blockers on quantal acetylcholine secretion indicates possible action of endogenous catecholamines on neuromuscular transmission. Elucidating the molecular mechanisms by which clinically utilized adrenomimetics and adrenoblockers regulate synaptic vesicle release at the motor axon terminal will lead to the creation of improved and safer sympathomimetics for the treatment of various neurodegenerative diseases with synaptic defects.

The F-Actin Binding Protein Cortactin Regulates the Dynamics of the Exocytotic Fusion Pore through its SH3 Domain.

Upon cell stimulation, the network of cortical actin filaments is rearranged to facilitate the neurosecretory process. This actin rearrangement includes both disruption of the preexisting actin network and de novo actin polymerization. However, the mechanism by which a Ca2+ signal elicits the formation of new actin filaments remains uncertain. Cortactin, an actin-binding protein that promotes actin polymerization in synergy with the nucleation promoting factor N-WASP, could play a key role in this mechanism. We addressed this hypothesis by analyzing de novo actin polymerization and exocytosis in bovine adrenal chromaffin cells expressing different cortactin or N-WASP domains, or cortactin mutants that fail to interact with proline-rich domain (PRD)-containing proteins, including N-WASP, or to be phosphorylated by Ca2+-dependent kinases, such as ERK1/2 and Src. Our results show that the activation of nicotinic receptors in chromaffin cells promotes cortactin translocation to the cell cortex, where it colocalizes with actin filaments. We further found that, in association with PRD-containing proteins, cortactin contributes to the Ca2+-dependent formation of F-actin, and regulates fusion pore dynamics and the number of exocytotic events induced by activation of nicotinic receptors. However, whereas the actions of cortactin on the fusion pore dynamics seems to depend on the availability of monomeric actin and its phosphorylation by ERK1/2 and Src kinases, cortactin regulates the extent of exocytosis by a mechanism independent of actin polymerization. Together our findings point out a role for cortactin as a critical modulator of actin filament formation and exocytosis in neuroendocrine cells.

Opioid and Opiate Immunoregulatory Processes.

The discovery of the ability of the nervous system to communicate through "public" circuits with other systems of the body is attributed to Ernst and Berta Scharrer, who described the neurosecretory process in 1928. Indeed, the immune system has been identified as another important neuroendocrine target tissue. Opioid peptides are involved in this communication (i.e., neuroimmune) and with that of autoimmunoregulation (communication between immunocytes). The significance of opioid neuropeptide involvement with the immune system is ascertained from the presence of novel δ, μ., and κ receptors on inflammatory cells that result in modulation of cellular activity after activation, as well as the presence of specific enzymatic degradation and regulation processes. In contrast to the relatively uniform antinociceptive action of opiate and opioid signal molecules in neural tissues, the presence of naturally occurring morphine in plasma and a novel μ3 opiate-specific receptor on inflammatory cells adds to the growing knowledge that opioid and opiate signal molecules may have antagonistic actions in select tissues. In examining various disorders (e.g., human immunodeficiency virus, substance abuse, parasitism, and the diffuse inflammatory response associated with surgery) evidence has also been found for the involvement of opiate/opioid signaling in prominent mechanisms. In addition, the presence of similar mechanisms in man and organisms 500 million years divergent in evolution bespeaks the importance of this family of signal molecules. The present review provides an overview of recent advances in the field of opiate and opioid immunoregulatory processes and speculates as to their significance in diverse biological systems.

Morphological and functional aspects of neurosecretory process in hypothalamic nuclei in heart failure

V.G. Papkov Hypothalamus studied in 18 men who died from injuries, acute coronary insufficiency and congestive heart failure. In the supraoptic (SOY) and paraventricular (PVN) nuclei determined percentage neurocyte that are in various phases of neurosecretory cycle. Calculated the index of the functional activity of the nuclei. Revealed an uneven change in the ratio neurocyte that are in various phases of neurosecretion and index decreased functional activity of SOY in various forms of heart failure. The index of the functional activity of the PVN in acute coronary insufficiency remains on the "control" value. The observed changes are regarded as a morphological indicator of the regulatory function of the hypothalamus.

Pannexin 1 channels: new actors in the regulation of catecholamine release from adrenal chromaffin cells.

Chromaffin cells of the adrenal gland medulla synthesize and store hormones and peptides, which are released into the blood circulation in response to stress. Among them, adrenaline is critical for the fight-or-flight response. This neurosecretory process is highly regulated and depends on cytosolic [Ca2+]. By forming channels at the plasma membrane, pannexin-1 (Panx1) is a protein involved in many physiological and pathological processes amplifying ATP release and/or Ca2+ signals. Here, we show that Panx1 is expressed in the adrenal gland where it plays a role by regulating the release of catecholamines. In fact, inhibitors of Panx1 channels, such as carbenoxolone (Cbx) and probenecid, reduced the secretory activity induced with the nicotinic agonist 1,1-dimethyl-4-phenyl-piperazinium (DMPP, 50 μM) in whole adrenal glands. A similar inhibitory effect was observed in single chromaffin cells using Cbx or 10Panx1 peptide, another Panx1 channel inhibitors. Given that the secretory response depends on cytosolic [Ca2+] and Panx1 channels are permeable to Ca2+, we studied the possible implication of Panx1 channels in the Ca2+ signaling occurring during the secretory process. In support of this possibility, Panx1 channel inhibitors significantly reduced the Ca2+ signals evoked by DMPP in single chromaffin cells. However, the Ca2+ signals induced by caffeine in the absence of extracellular Ca2+ was not affected by Panx1 channel inhibitors, suggesting that this mechanism does not involve Ca2+ release from the endoplasmic reticulum. Conversely, Panx1 inhibitors significantly blocked the DMPP-induce dye uptake, supporting the idea that Panx1 forms functional channels at the plasma membrane. These findings indicate that Panx1 channels participate in the control the Ca2+ signal that triggers the secretory response of adrenal chromaffin cells. This mechanism could have physiological implications during the response to stress.

Colocalization of FM1-43, Bassoon, and GnRH-1: GnRH-1 Release from Cell Bodies and Their Neuroprocesses

Pulsatile release of GnRH-1 is critical for reproductive function. However, the cellular mechanism of GnRH-1 neurosecretion is still elusive. In this study, we examined the neurosecretory process of GnRH-1 neurons using time-lapse image acquisition followed by immunocytochemistry with confocal microscopy. To monitor exocytotic processes, cultured GnRH-1 neurons derived from monkey embryos were labeled with the lipophilic dye, FM1-43, or its fixable form FM1-43Fx, in the presence or absence of depolarization signals, and changes in vesicles labeled with FM1-43 were analyzed. The results show FM1-43 was taken up into the cell and labeled puncta in the soma and neuroprocesses in the absence of depolarization signals, indicating that GnRH-1 neurons were spontaneously active. Depolarization of GnRH-1 neurons with high K+ or veratridine challenge increased the intensity and size of puncta in both soma and neuroprocesses, and the veratridine-induced changes in puncta were blocked by tetrodotoxin, indicating that changes in the puncta intensity and size reflect neurosecretory activity. Subsequent double immunocytochemistry for GnRH-1 and the synaptic vesicle marker, vesicle-associated membrane protein, demonstrated that the FM1-43Fx-labeled puncta were synaptic vesicles with the GnRH-1 peptide. Additional double immunocytochemistry for GnRH-1 and the marker of the neurosecretory active zone, Bassoon, indicated that the FM1-43Fx-labeled puncta were located at the sites of neurosecretory active zones in GnRH-1 neurons. These results suggest that GnRH-1 neurons have the capacity to release the peptide from the soma and dendrites. Collectively, we hypothesize that soma-dendritic release of the peptide may be a mechanism of synchronized activity among GnRH-1 neurons.

Presynaptic kainate and NMDA receptors are implicated in the modulation of GABA release from cortical and hippocampal nerve terminals

One of the pathways implicated in a fine-tuning control of synaptic transmission is activation of the receptors located at the presynaptic terminal. Here we investigated the intracellular events in rat brain cortical and hippocampal nerve terminals occurring under the activation of presynaptic glutamate receptors by exogenous glutamate and specific agonists of ionotropic receptors, NMDA and kainate. Involvement of synaptic vesicles in exocytotic process was assessed using [ 3H]GABA and pH-sensitive fluorescent dye acridine orange (AO). Glutamate as well as NMDA and kainate were revealed to induce [ 3H]GABA release that was not blocked by NO-711, a selective blocker of GABA transporters. AO-loaded nerve terminals responded to glutamate application by the development of a two-phase process. The first phase, a fluorescence transient completed in ∼1 min, was similar to the response to high K +. It was highly sensitive to extracellular Ca 2+ and was decreased in the presence of the NMDA receptor antagonist, MK-801. The second phase, a long-lasting process, was absolutely dependent on extracellular Na + and attenuated in the presence of CNQX, the kainate receptor antagonist. NMDA as well as kainate per se caused a rapid and abrupt neurosecretory process confirming that both glutamate receptors, NMDA and kainate, are involved in the control of neurotransmitter release. It could be suggested that at least two types ionotropic receptor are attributed to glutamate-induced two-phase process, which appears to reflect a rapid synchronous and a more prolonged asynchronous vesicle fusion.

Crustacean neuroendocrine systems and their signaling agents

Decapod crustaceans have long served as important models for the study of neuroendocrine signaling. For example, the process of neurosecretion was first formally demonstrated by using a member of this order. In this review, the major decapod neuroendocrine organs are described, as are their phylogenetic conservation and neurochemistry. In addition, recent advances in crustacean neurohormone discovery and tissue mapping are discussed, as are several recent advances in our understanding of hormonal control in this group of animals.

In vitro functionality of isolated embryonic hypothalamic vasopressinergic and oxytocinergic neurons: modulatory effects of brain-derived neurotrophic factor and angiotensin II

There are only a few studies on the ontogeny and differentiation process of the hypothalamic supraoptic-paraventriculo-neurohypophysial neurosecretory system. In vitro neuron survival improves if cells are of embryonic origin; however, surviving hypothalamic neurons in culture were found to express small and minimal amounts of arginine-vasopressin (AVP) and oxytocin (OT), respectively. The aim of this study was to develop a primary neuronal culture design applicable to the study of magnocellular hypothalamic system functionality. For this purpose, a primary neuronal culture was set up after mechanical dissociation of sterile hypothalamic blocks from 17-day-old Sprague-Dawley rat embryos (E17) of both sexes. Isolated hypothalamic cells were cultured with supplemented (B27)-NeuroBasal medium containing an agent inhibiting non-neuron cell proliferation. The neurosecretory process was characterized by detecting AVP and OT secreted into the medium on different days of culture. Data indicate that spontaneous AVP and OT release occurred in a culture day-dependent fashion, being maximal on day 13 for AVP, and on day 10 for OT. Interestingly, brain-derived neurotrophic factor (BDNF) and Angiotensin II (A II) were able to positively modulate neuropeptide output. Furthermore, on day 17 of culture, non-specific (high-KCl) and specific (Angiotensin II) stimuli were able to significantly (P<0.05) enhance the secretion of both neuropeptides over respective baselines. This study suggests that our experimental design is useful for the study of AVP- and OT-ergic neuron functionality and that BDNF and A II are positive modulators of embryonic hypothalamic cell development.

Crustacean neuropeptides.

Crustaceans have long been used for peptide research. For example, the process of neurosecretion was first formally demonstrated in the crustacean X-organ-sinus gland system, and the first fully characterized invertebrate neuropeptide was from a shrimp. Moreover, the crustacean stomatogastric and cardiac nervous systems have long served as models for understanding the general principles governing neural circuit functioning, including modulation by peptides. Here, we review the basic biology of crustacean neuropeptides, discuss methodologies currently driving their discovery, provide an overview of the known families, and summarize recent data on their control of physiology and behavior.

Exogenous Glutamate-Induced Modulation of Neurosecretory Process in Nerve Terminals Obtained from the Rat Brain

We studied intracellular processes in nerve terminals of neurons of the rat brain in response to application of exogenous glutamate. Using a рН-sensitive fluorescence probe, acridine orange (AO), and labeled gammaaminobutyric acid ([3Н]GABA), we estimated the effect of application of glutamate on the level of acidification of synaptic vesicles and also on the release of GABA from nerve terminals (synaptosomes) obtained from hippocampal tissue. Our experiments showed that glutamate in a dose-dependent manner stimulated the [3Н]GABA release from nerve terminals, and then we observed re-uptake of this neurotransmitter. A selective blocker of GABA transporters, NO-711, completely blocked the uptake of neurotransmitter but did not influence its release; this observation indicates that the glutamate-induced GABA release was from the vesicular, not cytosolic, pool. We confirmed that glutamate stimulates the process of exocytosis in experiments using AO, where we obtained data indicating that this process is two-phase. The first phase, which reflects probably calcium-induced exocytosis, looked like a “burst” of fluorescent signal typical of the response of synaptosomes to the action of KCl applied in depolarization concentration. Both phases of the response were completely blocked or significantly suppressed in calcium-free medium or in the presence of 25 μM Cd2+. The second (slow) phase of the response developed after a certain lag period and was characterized by a gradual increase in the intensity of fluorescent signal. This phase was completely dependent on the presence of sodium in the extracellular medium and completely blocked when sodium was replaced by choline or N-methyl-D-glucamine. We hypothesize that the second phase of the response can reflect either spontaneous unstimulated exocytosis or dissipation of the proton gradient in synaptic vesicles induced by the entry of Na+ into the nerve terminal.

Expression of the neurosecretory process in pc12 cells is governed by rest

The neurosecretory process is acquired during differentiation and can be lost en block by differentiated cells. To investigate the role of REST/NRSF, a transcription repressor, in the maintenance of the process we studied two PC12 clones, one wt and one defective, expressing low and high levels of endogenous RE-1 silencing transcription (factor) (REST), respectively. Stable transfection of constructs demonstrated that REST represses 10 genes coding for proteins of neurosecretory vesicles and their exocytosis, eight including and two lacking the REST-binding sequence, RE-1. Of these genes, those of chromogranins were strongly repressed by fewfold increases of REST, those of VAMP2 and syntaxin1a required much higher levels. Moreover, in wt cells transfected with an active construct the dense-core vesicles, still competent for regulated exocytosis, were much smaller, with lighter cores; in defective cells, the dominant-negative construct induced the rescue of many vesicle/exocytosis genes but not of those of chromogranins. Small dense-core vesicles, exocytized upon stimulation, were rescued when the construct-transfected defective cells were transfected also with chromograninA or treated with trichostatinA, a blocker of histone deacetylases. Our results identify REST, working by direct and indirect mechanisms, as the factor governing the maintenance of the neurosecretory process and the properties of dense-core vesicles in PC12 cells.

Imaging the neurosecretory process with FM1-43 in primate LHRH neurons

Imaging the neurosecretory process with FM1-43 in primate LHRH neurons

Distribution of synaptobrevin/VAMP 1 and 2 in rat brain

The synaptobrevin/vesicle-associated membrane protein (VAMP) family of proteins, which are essential for neurotransmitter release, are the vesicle donor soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor (SNARE) proteins first described in synaptic vesicles at nerve terminals. Two synaptobrevin/VAMP isoforms are involved in calcium-dependent synaptic vesicle exocytosis, synaptobrevin/VAMP 1 and synaptobrevin/VAMP 2. However, the functional significance of these two highly homologous isoforms remains to be elucidated. Here, we used immunohistochemical, immunofluorescence and confocal microscope techniques to localize the two synaptobrevin/VAMP isoforms in rat brain areas, particularly in nerve terminals. Our results show that the two isoforms are present in the rat central nervous system and that their expression overlaps in some areas. However, a distinct distribution pattern was detected. Synaptobrevin/VAMP 2 is the most abundant isoform in the rat brain and is widely distributed. Although synaptobrevin/VAMP 1 is less abundant, it is the main isoform in particular brain areas (e.g. zona incerta at the subthalamus or nerve terminals surrounding thalamic neurons). The colocalization of synaptophysin with synaptobrevin/VAMP 1 demonstrates the presence of this isoform in subsets of nerve terminals. These results indicate that each synaptic vesicle donor SNARE protein isoform could have a specialized role in the neurosecretory process.

Recruitment of synapses in the neurosecretory process during long-term facilitation at the lobster neuromuscular junction

We investigated long-term facilitation at the lobster neuromuscular synapse employing a combination of FM1-43 staining of synaptic vesicles, electron microscopy analysis, and electrical recordings of synaptic activity. Synaptic terminals were loaded with the fluorescent dye FM1-43 producing clusters of activity-dependent fluorescent spots. Electron microscopy analysis of synaptic ultrastructure suggested that fluorescent spots represent compartments of synaptic terminals filled with vesicles. Excitatory postsynaptic currents were recorded from the stained synaptic terminals using focal macropatch electrodes. Terminals were stained during the nerve stimulation at a low stimulation frequency (2, 5 or 10Hz) before and after long-term facilitation was elicited by high-frequency stimulation (20 or 30Hz for 5 min). We found that staining after long-term facilitation results in the appearance of new fluorescent spots, as well as in the increase in fluorescence of the spots that appeared before long-term facilitation. This increase in fluorescence accounted for the increase in quantal release. Activation of individual fluorescent spots was found to be non-uniform. In spite of overall increase in fluorescence, some synaptic compartments decreased their staining after long-term facilitation. Thus, our study demonstrates that long-term facilitation produces non-uniform activation of FM1-43 uptake in synaptic compartments that correlates with the increase in quantal neurosecretion.

Contributions of insect research toward our understanding of neurosecretion.

The process of neurosecretion is an important and widespread method of biological communication among animals. Although insects and vertebrates appear to be very different, neurosecretory mechanisms and the neuropeptides themselves are often the same. The gradual acceptance of neurosecretion as a biological phenomenon, largely as a result of research done with insects, is discussed.

Organization of afferent and efferent projections in the hypothalamic subiculum-supraoptic region system in the rat hypothalamus.

characteristic of the subiculum as a projection system is the fact that it also operates as an important integrative center for the limbic system as a whole. The neurons of this structure receive axons from pyramidal cells in all fields of the hippocampus, from neurons of the amygdala, and from neurons in the entorhinal cortex [4, 7]. Data on the role of this structure in regulating the neurosecretory process at the level of the neurosecretory nuclei of the hypothalamus and in organizing limbic-hypothalamic projections are very limited. However, it has been shown that damage to the ventral subiculum leads to spontaneous release of oxytocin, and also facilitates the milkreflex [8]; electrical stimulation of this structure results in inhibition of background spike activity in neurosecretory cells (NSC) in the supraoptic nucleus, identified by an antidromal stimulation method [1]. Additionally, morphological data indicate that the quantity of extrahypothalamic projections addressed directly by oxytocinergic and vasopressinergic cells of the supraoptic nucleus is small [9]. Direct projections of the ventral subiculum were seen only in the region adjacent to the nucleus. This region is currently being studied by many groups, using both morphological and electrophysiological approaches. It is perfectly possible that cells in this area may function as mediators, relaying the transmission of the inhibitory effects of the subiculum to NSC in the nucleus. In fact, the region adjacent to the supraoptic nucleus has been found to contain GABAergic neurons which could function as inhibitory interneurons [6]. It has also been demonstrated that electrical stimulation of the areas located dorsally and dorsolaterally to the supraoptic nucleus and in its immediate vicinity leads to activation or inhibition of neurons in this nucleus [5]. The aim of the present work was to use electrophysiological methods to investigate the involvement of neurons in the perinuclear zone in the transmission of the inhibitory influences of the subiculum on NSC of the supraoptic nucleus. Neuron spike activity was recorded extracellularly using glass microelectrodes. The margins of the nucleus were defined in terms of the detection of NSC, which were identified from the characteristics of their responses to stimulation of the peduncle of the hypophysis. Neuron recording was started 300-500/~m above the dorsal margin of the supraoptic nucleus. Additionally, neurons were recorded in parallel tracks lacking NSC, by repeated insertion of the same microelectrode, assuming that the electrode moved only minimally. The subiculum and the peduncle of the hypophysis were stimulated using concentric metal electrodes and monophasic square-wave currents. Stimulation was performed using occasionalsingle impulses at frequencies of 0.1-0.5 per sec. Electrodes were inserted using coordinates from a stereotaxic atlas of the rat brain [3] with subsequent checking of the position by histological investigation. Electrical stimulation of the subiculum produced activation in 47 % of the neurons recorded in the juxtanuclear region; 15% of cells showed reductions in the background spike frequency, while the remaining cells did not respond to the stimulus. Excitation and inhibition responses were mostly phasic, i.e., there was a small latent period and responses were of short duration (see Fig. I, A). The latent period varied from 4 to 40 msec in different neurons, and responses could consist of one or a series of action potentials. The data obtained here suggest that the projection of the subiculum to NSC in the supraoptic nucleus are not monosynaptic, but are at least oligosynaptic. The probability of seeing both activatory and inhibitory responses approached unity. There were no significant differences in the numbers of responding neurons or in the latent periods of their responses at different sites of cell recording (in tracks containing NSC or in parallel tracks).

GoProtein Does Not Regulate ATP-Stimulated [Ca2+]iElevation or Noradrenaline Release in PC12 Cells

The roles of Go, a heterotrimeric GTP binding (G) protein with a 40-kDa α subunit and which is localized predominantly in neuronal cells, in exocytosis have been discussed recently. PC12 pheochromocytoma cell line is a convenient model in which to study the Ca2+-dependent mechanisms of the neurosecretory process. The stimulation of ATP receptors or addition of KCl stimulated an elevation of cytosolic free Ca2+concentration ([Ca2+]i) and [3H]noradrenaline (NA) release in PC12 cells. In this study, we investigated the roles of Goin ATP- and KCl-stimulated reactions. Nerve growth factor treatment for 2 days and transfection of PC12 cells with cDNA of subunit of Go(Goα) had no effect on ATP-stimulated [3H]NA release, although both treatments increased levels of the Goαand its trimeric form by about twofold over those in unstimulated cells. The [Ca2+]irise induced by ATP in NGF-treated cells was similar to that in control cells, although the maximal response was slightly smaller. Cholera toxin treatment for 2 days inhibited ATP- and KCl-stimulated NA release, although this treatment caused an approximately twofold increase in the level of Go. Pertussis toxin treatment, which ADP ribosylated over 90% of endogenous G proteins such as Go, had no effect on ATP-stimulated reactions. These findings show that Godoes not directly regulate ATP-stimulated Ca2+channels or the NA release process in PC12 cells. Cholera toxin-sensitive protein(s) may regulate exocytosis.

Opioid and opiate immunoregulatory processes.

The discovery of the ability of the nervous system to communicate through "public" circuits with other systems of the body is attributed to Ernst and Berta Scharrer, who described the neurosecretory process in 1928. Indeed, the immune system has been identified as another important neuroendocrine target tissue. Opioid peptides are involved in this communication (i.e., neuroimmune) and with that of autoimmunoregulation (communication between immunocytes). The significance of opioid neuropeptide involvement with the immune system is ascertained from the presence of novel delta, mu, and kappa receptors on inflammatory cells that result in modulation of cellular activity after activation, as well as the presence of specific enzymatic degradation and regulation processes. In contrast to the relatively uniform antinociceptive action of opiate and opioid signal molecules in neural tissues, the presence of naturally occurring morphine in plasma and a novel mu3, opiate-specific receptor on inflammatory cells adds to the growing knowledge that opioid and opiate signal molecules may have antagonistic actions in select tissues. In examining various disorders (e.g., human immunodeficiency virus, substance abuse, parasitism, and the diffuse inflammatory response associated with surgery) evidence has also been found for the involvement of opiate/opioid signaling in prominent mechanisms. In addition, the presence of similar mechanisms in man and organisms 500 million years divergent in evolution bespeaks the importance of this family of signal molecules. The present review provides an overview of recent advances in the field of opiate and opioid immunoregulatory processes and speculates as to their significance in diverse biological systems.

Exposure to Highsalinitymediumandneurosecretion in Theanteromedial Cells Ofthesupraesophageal Ganglion Ofthe Fresh-watershrimpMacrobrachiumolfersii (Decapoda)

ABSTRACT Neurofactors released from neurosecretory cells associated with the ganglia of the central nervous system are known to participate in the endocrine mediation of osmotic and ionic regulation in decapod Crustacea. The present study examines the effects of differential exposure (3, 6, 12, or 24 h) to high salinity media (HSM, sea water of 14, 21, or 28‰ salinity) on the ultrastructure of the anteromedial cells located within the protocerebral region of the supraesophagcal ganglion of Macrobrachium olfersii (Wiegmann), a euryhaline, fresh-water shrimp. Analysis of thin sections and freeze-fracture replicas reveals that the cells of the anteromedial group are typically neurosecretory in nature, exhibiting a large nucleus, a prominent, single nucleolus, well-developed, rough endoplasmic reticulum, numerous, large Golgi complexes, abundant mitochondria, and free and polyribosomes. In shrimps maintained in river water (0‰ salinity), this cellular machinery, specific for the production and packaging of polypeptides, does not appear to be activated, as indicated by the virtual absence of neurosecretory granules. After exposure of the shrimps to HSM, particularly to 21‰ salinity, for up to 6 h, there is a conspicuous increase in the number of dense-cored, neurosecretory granules of 72.4 ± 6.7 nm diameter, both in the cytoplasm and around the Golgi complexes of these cells. Multivesicular and multilamellar bodies also increase in number, indicating membrane recycling. In shrimps exposed to HSM for longer periods (12 h or more), the ultrastructure of the anteromedial cells is essentially similar to that of control shrimps maintained in fresh water. These data demonstrate that exposure to high salinity media activates a neurosecretory process in the anteromedial cells of the protocerebrum of M. olfersii. The release of these neurosecretory products may be involved in osmotic and ionic regulation in this fresh-water shrimp.