Involvement of protein kinase C in the galanin-induced antinociception in the brain of rats

Previous study in our laboratory demonstrates that microinjection of galanin into the arcuate nucleus of hypothalamus produced antinociceptive effects in rats. In the present study we investigated the involvement of protein kinase C (PKC) and PKC signaling pathways in the galanin-induced antinociception in the brain of rats. Intracerebroventricular injection of galanin produced antinociceptive effects in rats tested by hot-plate and Randall Selitto test. Interestingly, the galanin-induced antinociception was significantly attenuated by intracerebroventricular injection of the PKC inhibitor chelerythrine, indicating an involvement of PKC in the galanin-induced antinociception in rats. Taken together, the results demonstrate that galanin induces antinociceptive effects in the rat brain, and PKC is involved in the galanin-induced antinociception in the brain of rats.     

Antinociceptive effects of intracerebroventricular injection of the galanin receptor 1 agonist M 617 in rats

Previous studies in our laboratory demonstrated that galanin and its receptors play important roles in nociceptive modulation in the central nervous system. The present study was performed to explore the antinociceptive effects of the galanin receptor 1 agonist M 617 in the central nervous system of rats. Intracerebroventricular injection of 0.1nmol, 0.5nmol, 1nmol or 2nmol of M 617 induced dose-dependent increases in hindpaw withdrawal latencies (HWLs) to noxious thermal and mechanical stimulations in rats. Furthermore, both intracerebroventricular injection of M 617 and galanin induced significant increases in HWLs in rats. Interestingly, there were no significant differences between the antinociceptive effects induced by M 617 and galanin, indicating that galanin receptor 1 plays main roles in galanin-induced antinociceptive effects in the brain of rats.     

Antinociceptive effects induced by injection of the galanin receptor 1 agonist M617 into central nucleus of amygdala in rats

The present study was performed to explore the antinociceptive effects of M617, a selective galanin receptor 1 agonist, in the central nucleus of amygdala (CeA) of rats. Intra-CeA injection of 0.1 nmol, 0.5 nmol and 1 nmol of M617 induced dose-dependent increases in hindpaw withdrawal latencies (HWLs) to noxious thermal and mechanical stimulations in rats. Furthermore, rats received intra-CeA administration of M617 and galanin. The HWL to noxious thermal and mechanical stimulations increased markedly, and there were no significant differences in HWLs of rats received intra-CeA administration of M617 and galanin. The results demonstrated that intra-CeA injection of M617 induced significant antinociceptive effects in CeA of rats, indicating that galanin receptor 1 may be involved in M617-induced antinociception in the CeA of rats.     

Centrally administered galanin inhibits osmotically stimulated arginine vasopressin release in conscious rats.

The effect of centrally administered galanin on arginine vasopressin (AVP) release was investigated in conscious rats. Intracerebroventricular injection of porcine galanin suppressed hypertonic saline-induced increase in plasma AVP in a dose-dependent manner (12.5-100 pmol/rat) at 10 min after the injection. Pretreatment with subcutaneous injection of naloxone (1 mg/100 g b.wt.) partially blocked the galanin-induced effect on plasma AVP. These results suggest that central galanin inhibits osmotically stimulated AVP release and endogenous opioids are, at least in part, involved in the mechanism.     

Analysis of the galanin-induced decrease in membrane excitability in mudpuppy parasympathetic neurons.

Previously, we showed that the neuropeptide galanin hyperpolarizes and decreases membrane excitability of mudpuppy parasympathetic neurons [Konopka L. M., McKeon T. W. and Parsons R. L. (1989) J. Physiol. 410, 107-122]. We also demonstrated that membrane excitability remains depressed when the agonist-induced potential change is negated electrotonically. We hypothesized that galanin inhibits the membrane conductances associated with spike generation. However, we cannot rule out the possibility that the decreased excitability is due to a galanin-induced increase in membrane potassium conductance which reduces the effectiveness of subsequent depolarizing stimuli. Therefore, in the present study we tested, with the galanin-induced hyperpolarization negated, whether the galanin-induced increased membrane potassium conductance was responsible for the decreased excitability. The results showed that the galanin-induced decreased excitability was not dependent on the peak amplitude of the galanin-induced hyperpolarization. Furthermore, the decreased excitability occurred in cells in which there was no measurable galanin-induced hyperpolarization. Moreover, in most cells the galanin-induced decrease in input resistance, measured at the peak of the hyperpolarization (3-25 mV), was less than 15% and when the hyperpolarization was negated electronically, the decrease was even less (approximately 2%). These results indicated that when the hyperpolarization was negated, the galanin-induced increase in potassium conductance was not responsible for the decreased excitability. In preparations pretreated with 5 mM tetraethylammonium, galanin decreased excitability which indicated that a galanin-induced decrease in the calcium-dependent potassium current was not necessary for the decreased excitability. Galanin also decreased excitability in preparations exposed to either 1-3 microM tetrodotoxin or 100-200 microM cadmium. Following galanin application, the threshold for initiation of tetrodotoxin-insensitive spikes was shifted to more positive membrane potentials. Galanin also decreased the amplitude and hyperpolarizing afterpotential of barium spikes in the absence of any agonist-induced hyperpolarization. These observations confirmed that galanin decreased the voltage-dependent calcium conductance. In the present study, we showed that when the hyperpolarization was negated, galanin decreased excitability by shifting the threshold for spike generation regardless of whether voltage-dependent sodium or calcium currents were primarily responsible for the depolarizing component of the action potential.     

Post-infarct cardiac sympathetic hyperactivity regulates galanin expression

The neuropeptide galanin is elevated in the cardiac sympathetic innervation after myocardial infarction (MI). Galanin inhibits vagal transmission and may support the regeneration of sympathetic nerves, thereby contributing to the development of arrhythmia and sudden cardiac death after MI. The reason for increased galanin production in sympathetic neurons after myocardial infarction is not known. Cardiac sympathetic neurons are activated chronically after cardiac ischemia–reperfusion, and activation of sympathetic neurons in culture stimulates galanin expression. Therefore, we tested the hypothesis that increased sympathetic nerve activity stimulates galanin expression in cardiac sympathetic neurons after myocardial infarction. To test this hypothesis we used TGR(ASrAOGEN) transgenic rats, which lack brain angiotensinogen and do not exhibit post-infarct sympathetic hyperactivity. Hearts and stellate ganglia were collected 1 week after ischemia–reperfusion. Galanin mRNA was quantified by real-time PCR and peptide content was assayed by enzyme-linked immunosorbent assay. Galanin mRNA increased approximately 3-fold after MI in cardiac sympathetic neurons of both genotypes compared to unoperated and sham controls. Left ventricular galanin content, however, increased after MI only in Sprague–Dawley rats and not in AOGEN rats. These data suggest that post-infarct cardiac sympathetic hyperactivity stimulates galanin peptide production but is not required for increased galanin mRNA expression.     

Galanin: effects on basal and stimulated insulin and glucagon secretion in the mouse

Galanin was recently demonstrated to be a neuropeptide in intrapancreatic nerves. In this study, the effects of galanin on basal and stimulated insulin and glucagon secretion in the mouse were investigated. Galanin, injected intravenously at dose levels ranging from 0.53 to 8.5 nmol kg-1, markedly lowered basal plasma insulin levels and transiently increased basal plasma glucagon levels. Furthermore, galanin induced hyperglycaemia: plasma glucose levels were 11 +/- 0.2 mmol l-1 2 min after injection of galanin (4.25 nmol kg-1) compared with 9.3 +/- 0.3 mmol-1 in controls (P less than 0.001). Galanin also impaired the plasma insulin response to either glucose or the cholinergic agonist carbachol. Thus, galanin (4.25 nmol kg-1) inhibited the plasma insulin response to glucose by 65% (P less than 0.001), and that to carbachol by 85% (P less than 0.001). Moreover, glucose abolished the galanin-induced plasma glucagon response. Also, galanin and carbachol exerted additive stimulatory effects on glucagon levels. It is concluded from this study in mice that galanin inhibits basal and stimulated insulin secretion, stimulates glucagon secretion, and induces hyperglycaemia. It is suggested that the intrapancreatic neuropeptide galanin is of importance in the regulation of both insulin and glucagon secretion.     

An ERP study on the time course of facial trustworthiness appraisal

The neuropeptide galanin has been recognized as a possible neurotransmitter/neuromodulator, and in addition has been implicated in anxiety- and depression-related behaviors. The present study demonstrates increased locomotion and rearing after galanin (0.3 mg/kg) that was given intraperitoneally (i.p.) to intact Wistar rats which were tested 1 h later in the open field (OF). These effects, which suggest an anxiolytic-like action, were blocked by i.p. administered peptidic galanin antagonist M40. Further, the locomotion increase caused by galanin and the inhibitory effect of M40 persisted for 48 h without additional treatment. Rats exposed to restraint stress (lasting 60 min) for three consecutive days and tested 1 h after stress termination exhibited reduced locomotion and exploration in the OF. Galanin (0.3 and 1.0 mg/kg) given immediately after each stress exposure prevented the decrease of locomotion and exploration induced by stress in all trials. When the test was repeated 6 days later without stress and galanin treatment the reduction of locomotion produced by stress persisted; the anti-stress behavioral effects of both galanin doses were also present. Testing performed on the 12th day after the last stress and galanin treatment with 0.3 mg/kg revealed an increased locomotion compared with unstressed and stress-exposed rats. Our results demonstrate that behavioral effects of the peptide galanin are evident even after i.p. administration. These results also suggest that galanin elicits stress-modulatory action, and support the notion that the galaninergic system may serve as a drug target in stress-related conditions.     

Distribution of galanin in bone and joint tissues

The article describes the distribution of galanin in normal bone and joint tissues. Periosteum, cortical bone, bone marrow, and synovial membrane of normal rats were analyzed. Immunoelectron microscopy (iEM) was used to analyze the distribution of galanin, and radioimmunoassay (RIA) was used to determine its concentration. Immunoelectron microscopy showed that galanin is abundant in nerve fibers and endothelial cells in the periosteum and also in macrophage-like-cells and nerve fibers of the synovial membrane. The concentration of galanin measured by RIA showed the highest concentration in bone marrow, followed by periosteum and cortical bone. This study demonstrates that galanin is present and can be quantified in different compartments of bone and joint tissues and illustrates the possible role of galanin under physiological conditions.     

Characteristics of the galanin-induced depolarization of mudpuppy parasympathetic postganglionic neurons

The depolarization of mudpuppy parasympathetic cardiac neurons, produced by pressure application of the neuropeptide galanin, has been characterized. The amplitude and duration of the depolarization were dependent on the duration of the galanin application. The amplitude of the depolarization increased with hyperpolarization and the reversal potential determined by extrapolation was approximately +10 mV. The amplitude and time course of the galanin-induced depolarization were not changed by substitution of either manganese or magnesium for extracellular calcium, but were decreased by exposure to 50-100 microM d-tubocurarine. It is proposed that the galanin-induced depolarization results from a receptor-activated, nonselective, cation channel; and further, that the receptor initiating depolarization differs from the galanin receptor mediating hyperpolarizing responses in these parasympathetic neurons.     

Involvements of mu- and kappa-opioid receptors in morphine-induced antinociception in the nucleus accumbens of rats

It is well known that there are three types of opioid receptors, mu- (MOR), delta- (DOR), and kappa-opioid receptor (KOR) in the central nervous system. The present study investigated the involvement of opioid receptors in morphine-induced antinociception in the nucleus accumbens (NAc) of rats. The hindpaw withdrawal latencies to thermal and mechanical stimulation increased markedly after intra-NAc administration of morphine. The antinociceptive effects induced by morphine were dose-dependently inhibited by intra-NAc administration of the non-selective opioid receptor antagonist naloxone. Furthermore, the morphine-induced antinociception was significantly attenuated by subsequent intra-NAc injection of the MOR antagonist beta-funaltrexamine or the KOR antagonist nor-binaltorphimine, but not the DOR antagonist naltrindole. The results indicate that MOR and KOR, but not DOR are involved in the morphine-induced antinociception in the NAc of rats.     

Antinociceptive role of galanin in periaqueductal grey of rats with experimentally induced mononeuropathy

The present study was performed in rats with experimentally induced mononeuropathy after left common sciatic nerve ligation. The hindpaw withdrawal latencies to thermal and mechanical stimulation increased significantly after intra-periaqueductal grey injection of 2 or 3nmol, but not 1nmol of galanin in rats with mononeuropathy. Intraperitoneal administration of 4.5mg/kg morphine induced significant increases in hindpaw withdrawal latencies to both noxious stimulation, which were attenuated by following intra-periaqueductal grey injection of 2nmol of the galanin antagonist galantide. Furthermore, the antinociceptive effect induced by intra-periaqueductal grey injection of 26.6nmol of morphine was attenuated significantly by following intra-periaqueductal gray administration of 2nmol of galantide.The results demonstrated that in periaqueductal grey galanin plays an antinociceptive role in rats with mononeuropathy and galanin is involved in the mechanisms of opioid-induced antinociception.     

Estrogen induction of galanin synthesis in the rat anterior pituitary gland demonstrated by in situ hybridization and immunohistochemistry

The distribution of galanin messenger (mRNA) and galanin-like immunoreactivity (Gal-LI) in the anterior and posterior pituitaries of control and estrogen-implanted female rats was determined by in situ hybridization and immunohistochemical methods. In control ovariectomized animals galanin mRNA was undetectable in the posterior, intermediate and anterior pituitary. However, 4 days after implantation with 10 mg of diethylstilbestrol, galanin mRNA was clearly present in the anterior pituitary, but not in the posterior or intermediate lobe. By immunohistochemistry Gal-LI was readily visualized and detected in the posterior lobe, but clearly undetectable in cells of the intermediate and anterior pituitary lobes of control animals. After estrogen administration numerous cells exhibiting intense Gal-LI were evident in the anterior lobe, while Gal-LI remained unchanged in the intermediate and posterior lobes. These results indicate that in control animals galanin is stored, but not synthesized, in the posterior pituitary and that after estrogen administration galanin production is substantially increased in the anterior pituitary. We conclude that the expression of galanin in the anterior pituitary is regulated by estrogen and suggest that galanin may be a pituitary hormone.     

Effect of M35, a neuropeptide galanin antagonist on glucose uptake translated by glucose transporter 4 in trained rat skeletal muscle

In this study, we used M35, a galanin antagonist to explore the effect of an increase in galanin release induced by exercise on glucose transporter 4 (GLUT4) content and function. The rats tested were divided into four groups: rats from sedentary and trained drug groups were injected by M35, 5 times per week during four weeks. Rats from sedentary and trained control groups by 0.1 mol/l citrate buffer. The rats from both exercise groups swam after each injection. The results showed that M35 significantly decreased glucose infusing speeds in euglycemic–hyperinsulinemic clamp tests. M35 treatment elevated plasma insulin levels in both drug groups. And the insulin levels in both drug groups were higher also than that after experiments in each control group respectively. The four weeks swimming enhanced the plasma galanin contents. The galanin levels after experiments in both exercise groups were higher than that in each sedentary control group respectively too. The GLUT4 densities were attenuated by M35 at plasma membranes and total cell membranes. The change ratios of GLUT4 immunoreaction at plasma membranes to total cell membranes were lower in both drug groups compared to each control group. Those results suggest that endogenous galanin has an important attribute to elevate the insulin sensitivity by increasing GLUT4 contents and promoting GLUT4 transportation from intracellular membranes to plasma membranes in muscle cells. Galanin is an important hormone to elevate insulin sensitivity in rest and exercise conditions.     

Orexin-mediated feeding behavior involves both leptin-sensitive and -insensitive pathways

Orexin-A and -B are neuropeptides that are implicated in the regulation of vigilance states and energy homeostasis. Orexins are specifically produced by neurons located within the lateral hypothalamic area (LHA), a region implicated in the regulation of feeding behavior. Here, we examined the functional interactions between orexins and anorectic factors [leptin, alpha-melanocyte-stimulating hormone (alpha-MSH) and glucagon-like peptide-1 (GLP-1)] in rats. Intracerebroventricular injection of orexin-A (10 nmol) potently augmented food intake in rats. Neuropeptide Y (NPY) (0.3 nmol) and galanin (3 nmol) also induced a transient increase in food intake. Both NPY- and galanin-induced feeding behaviors were completely inhibited by preadministration of leptin (3 microg), while the same or a higher dose (10 microg) of leptin only partially inhibited orexin-A or -B-induced increase of food intake. Preadministration of anorectic peptides (alpha-MSH and GLP-1), which are shown to be regulated by leptin, abolished NPY-induced feeding; however, orexin-induced feeding was only partially inhibited by these anorectic peptides. These observations suggest that NPY- and galanin-induced increases of feeding involve a leptin-sensitive pathway, while orexin-induced feeding involves both leptin-sensitive and -insensitive pathways.     

Galanin stimulation of feeding is blocked by the addition of a response requirement

Administration of the neuropeptide galanin increases food intake in laboratory rats and mice, however this increase has only been observed under conditions of free-feeding. As there is a growing distinction between consummatory and instrumental behavior, we assessed whether galanin would differentially affect food consumption when food was freely available or when the same food was response-contingent. We also tested whether food restriction would interact with galanin's effect on food consumption in either condition. As in previous studies, galanin significantly increased food consumption under free access conditions. However, when food was contingent upon lever-pressing, galanin had no such stimulatory effect. Food-restriction increased basal feeding levels in both tasks but there was no interaction between food restriction and free or response-contingent food intake. These results demonstrate that galanin-induced feeding stimulation is limited to conditions of free access and support the theoretical distinction between consummatory and instrumental behavior.     

Galanin-induced hyperpolarization of mudpuppy neurons is calcium dependent.

The calcium dependence of the hyperpolarization produced by galanin has been studied in parasympathetic postganglionic neurons of the mudpuppy. Necturus maculosus. The galanin-induced hyperpolarization, but not the bethanechol-induced hyperpolarization, was reduced when manganese or magnesium were substituted for external calcium. Also, the galanin-induced, but not the bethanechol-induced hyperpolarization, was decreased in the presence of cadmium or nitrendipine. Because the inhibition of the galanin-induced hyperpolarization developed gradually and reversed slowly in the absence of extracellular calcium or presence of the voltage-gated calcium channel blockers, it is suggested that an intracellular pool rather than an extracellular pool of calcium is involved in the generation of the galanin-induced hyperpolarization.     

Voluntary exercise and clomipramine treatment elevate prepro-galanin mRNA levels in the locus coeruleus in rats

Exercise exerts antidepressant effects in humans and rodent models of affective disorders. These effects may be mediated by the upregulation of endogenous factors that exert antidepressant actions. The physiological functions and behavioral actions of the neuropeptide galanin (GAL) suggest antidepressant activity. Previous studies have shown that various modes of exercise elevate GAL gene expression in the locus coeruleus (LC) in rats. The present experiments examined the interaction between voluntary exercise and antidepressant pharmacotherapy. Male Sprague–Dawley rats were provided access to activity wheels (exercise condition) or inoperative wheels (sedentary condition) for 28 days. Rats in each group were injected with clomipramine (10 mg/kg/day) or vehicle throughout this period (for 3 weeks). Prepro-GAL mRNA in the LC was measured by in situ hybridization histochemistry. Exercise and clomipramine treatment significantly elevated GAL gene expression, though prepro-GAL mRNA levels in rats receiving both interventions did not differ from sedentary controls that received vehicle. Prepro-GAL mRNA levels were significantly correlated with running distance. The results further implicate a role for GAL in the antidepressant effects of exercise and pharmacotherapy, though the mechanisms through which these treatments influence GAL gene expression appear to differ significantly.     

Ethanol intake increases galanin mRNA in the hypothalamus and withdrawal decreases it

Alcoholism can be viewed as a motivational disorder that results from alterations in brain systems for ingestive behavior. Therefore, it was hypothesized that alcohol intake might alter the expression of hypothalamic peptides that stimulate feeding. Earlier studies showed that hypothalamic injection of the feeding-stimulatory peptide, galanin (GAL), increases the release of dopamine (DA) in the nucleus accumbens (NAc), as does systemic alcohol, leading to a focus on GAL. Results of this study demonstrate the following: (1). Ethanol, injected daily (0.8 g/kg 10% v/v) for 7 days in male rats, markedly increased the expression of GAL but not of neuropeptide Y (NPY). This occurred in specific hypothalamic nuclei, namely the dorsomedial nucleus (DMN), paraventricular nucleus (PVN) and perifornical lateral hypothalamus (PLH). (2). Rats induced to drink ethanol ad libitum, by gradually increasing the concentration from 1% to 9% v/v without adding sugar or flavoring, exhibited a similar stimulation of GAL mRNA in the PVN and GAL immunoreactivity in the DMN and PVN. (3). Rats given increasing ethanol concentrations, with 12 h access starting 4 h into the dark cycle, had a mean blood alcohol concentration of 18 mg/dl and exhibited a similar increase in GAL expression in the DMN and PVN. (4) Withdrawal from the opioid effects of 9% ethanol, produced by injection of naloxone (3 mg/kg sc), reversed this ethanol effect by significantly reducing GAL expression in the DMN and PLH below baseline levels. These studies suggest a possible role for hypothalamic GAL in alcohol abuse.     

Effects of intraventricular injections of galanin on neuroendocrine functions in the male rat. Possible involvement of hypothalamic catecholamine neuronal systems

Galanin-catecholamine interactions have been analysed within the hypothalamus and the anteromedial frontal cortex of male rats by means of quantitative histofluorimetrical and biochemical measurements of catecholamine fluorescence in discrete catecholamine nerve terminal systems and measurements of serum levels of adenohypophyseal hormones and corticosterone using radio-immunoassay determinations. 125I-galanin binding sites were analysed and related to the distribution of galanin-immunoreactive neuronal structures in the median eminence and paraventricular hypothalamic nucleus. The results show that intraventricular injections of galanin in the awake and unrestrained male rat produce rapid increases of prolactin and growth hormone secretion but no effects on serum luteinizing hormone, thyroid stimulating hormone or on corticosterone levels. These changes in neuroendocrine function were associated with a selective reduction of the catecholamine stores in the medial palisade zone of the median eminence at the 20 min time interval. 125I-galanin binding sites were found throughout the hypothalamus including the median eminence and the magnocellular part of the paraventricular hypothalamic nucleus with a good correspondence with galanin immuno-reactivity. It is suggested that the enhancement of prolactin secretion induced by galanin involves an interaction between galanin and dopamine in the medial palisade zone leading to a reduced synthesis and/or release of dopamine and thus to a reduced prolactin inhibitory activity and to increases in prolactin secretion. A possible involvement of hypothalamic catecholamines in the galanin-induced changes of growth hormone secretion remains to be established.