Neurons In Paragigantocellularis Research Articles

Prolonged morphine exposure during adolescence alters the responses of lateral paragigantocellularis neurons to naloxone in adult morphine\xa0dependent rats

IntroductionAdolescence is a critical period in brain development, and it is characterized by persistent maturational alterations in the function of central nervous system. In this respect, many studies show the non-medical use of opioid drugs by adolescents. Although this issue has rather widely been addressed during the last decade, cellular mechanisms through which adolescent opioid exposure may induce long-lasting effects are not duly understood. The present study examined the effect of adolescent morphine exposure on neuronal responses of lateral paragigantocellularis nucleus to naloxone in adult morphine-dependent rats.MethodsAdolescent male Wistar rats (31 days old) received increasing doses of morphine (from 2.5 to 25 mg/kg, twice daily, s.c.) for 10 days. Control subjects were injected saline with the same protocol. After a drug-free interval (20 days), animals were rendered dependent on morphine during 10 days (10 mg/kg, s.c., twice daily). Then, extracellular single-unit recording was performed to investigate neural response of LPGi to naloxone in adult morphine-dependent rats.ResultsResults indicated that adolescent morphine treatment increases the number of excitatory responses to naloxone, enhances the baseline activity and alters the pattern of firing in neurons with excitatory responses in adult morphine-dependent rats. Moreover, the intensity of excitatory responses is reduced following the early life drug intake.ConclusionIt seems that prolonged opioid exposure during adolescence induces long-lasting neurobiological changes in LPGi responsiveness to future opioid withdrawal challenges.

Early life maternal deprivation attenuates morphine induced inhibition in lateral paragigantocellularis neurons in adult rats

Early life stress can serve as one of the principle sources leading to individual differences in confronting challenges throughout the lifetime. Maternal deprivation (MD), a model of early life stress, can cause persistent alterations in brain function, and it may constitute a risk factor for later incidence of drug addiction. It is becoming more apparent that early life MD predisposes opiate abuse in adulthood. Although several behavioral and molecular studies have addressed this issue, changes in electrophysiological features of the neurons are yet to be understood. The lateral paragigantocellularis (LPGi) nucleus, which participates in the mediation of opiate dependence and withdrawal, may be susceptible to modifications following MD. This study sought to find whether early life MD can alter the discharge activity of LPGi neurons and their response to acute morphine administration in adult rats.Male Wistar rats experienced MD on postnatal days (PNDs) 1–14 for three h per day. Afterward, they were left undisturbed until PND 70, during which the extracellular activities of LPGi neurons were recorded in anesthetized animals at baseline and in response to acute morphine.In both MD and control groups, acute morphine administration induced heterogeneous (excitatory, inhibitory, and no effect) responses in LPGi neurons. At baseline recording, the interspike interval variability of the LPGi neurons was attenuated in both inhibitory and excitatory responses in animals with the history of MD. The extent of morphine-induced discharge inhibition was also lower in deprived animals compared to the control group.These findings suggest that early life MD induces long-term alterations in LPGi neuronal activity in response to acute administration of morphine. Therefore, the MD may alter the vulnerability to develop opiate abuse in adulthood.

Direct Facilitatory Role of Paragigantocellularis Neurons in Opiate Withdrawal-Induced Hyperactivity of Rat Locus Coeruleus Neurons: An In Vitro Study

Studies have shown that following opiate withdrawal, the spontaneous discharge rate of locus coeruleus (LC) neurons remarkably increases. Combination of intrinsic mechanisms with extrinsic excitatory modulations mediates the withdrawal-induced hyperactivity of LC neurons. The nucleus paragigantocellularis (PGi) provides the main excitatory inputs to LC and plays a pivotal role in opiate withdrawal. In the present study the direct facilitatory role of PGi on opiate withdrawal-induced hyperactivity of LC neurons was investigated using a newly developed brain slice, containing both LC and PGi. HRP retrograde neuronal tracing was used to verify the existence of both LC and PGi neurons in the developed slice. The spontaneous discharge rate (SDR), resting membrane potential (RMP) and spontaneous excitatory post-synaptic currents (sEPSCs) were recorded in LC neurons using whole cell patch clamp recording. Results showed that the net SDR and the net RMP of LC neurons in slices containing both LC and PGi neurons are significantly higher than slices lacking intact (uncut) PGi inputs. Also, the frequency of sEPSCs in those LC neurons receiving PGi inputs significantly increased compared to the slices containing no intact PGi inputs. Altogether, our results propose that increase in PGi-mediated excitatory transmission might facilitate the opiate withdrawal-induced hyperactivity of LC neurons.

The effect of morphine on some electrophysiological parameters of paragigantocellularis and locus coeruleus nuclei interconnections

As one of the most important diffused brain modulatory systems, the nucleus locus coeruleus (LC) receives most of its afferents from the nucleus paragigantocellularis (PGi) and plays a major role in the control of drug dependence and some emotional and exciting states. For detailed investigation of the effect of morphine on relationship between these two brain stem nuclei, the activity of the rat (Sprague-Dawley) LC neurons was examined after being dependent on morphine in comparison with independent group. The activity of the nucleus LC neurons with and without the activity of nucleus PGi neurons was recorded using extracellular single unit recording technique. Lidocaine microinjection (1 ml, in 1-2 min, 2.5%) was used for inactivation of PGi. The PSTH (Peri-Stimulus Time Histogram) of the neuronal activity showed a decrease in neuronal firing rate of LC by 85.8±1.7% in morphine-dependent group (from 2.95 spike/sec to 0.64 spike/sec, n = 4) and by 48.3±4.3% (from 3.5 spike/sec to 1.94 spike/sec, n = 6) for independent one. The difference was significant (P

Discharge activities of neurons in the nucleus paragigantocellularis during the development of morphine tolerance and dependence: a single unit study in chronically implanted rats.

The nucleus paragigantocellularis (PGi) has been proposed to play a role in opiate dependence/withdrawal. In the present study, we examined the discharge activity of PGi neurons before and after the development of morphine tolerance/dependence in rats. A multi-wire electrode was chronically implanted in the PGi, which allowed us to monitor the effects of both acute and chronic morphine treatments on the activity of PGi neurons recorded from the same site. We found that acute morphine excited, inhibited or had no effect on 36%, 35% or 29% of PGi neurons (N=556), respectively. After 3 days of continuous morphine infusion, which led to morphine tolerance/dependence, the firing rates of both excitatory and inhibitory PGi neurons returned to pre-morphine treatment levels, indicating that the PGi neurons developed tolerance to both excitatory and inhibitory effects of morphine. Naltrexone-precipitated withdrawal from chronic morphine treatment also induced heterogeneous responses in the PGi. On a site-by-site basis, most of the sites that showed excitatory response to acute morphine exhibited inhibitory response during withdrawal, and all the sites that showed inhibitory response to acute morphine exhibited excitatory response during withdrawal. Correlation analysis further quantitatively showed that PGi neurons' responses to acute morphine and that during withdrawal were inversely correlated with a correlation coefficient of 0.73, suggesting that adaptations in the PGi during the development of morphine dependence share common neural mechanisms with the acute effect of morphine. These results provide new insights into the role of the PGi in the development of morphine tolerance/dependence.

Divergent projections of catecholaminergic neurons in the nucleus of the solitary tract to limbic forebrain and medullary autonomic brain regions

The nucleus of the solitary tract (NTS) is a critical structure involved in coordinating autonomic and visceral activities. Previous independent studies have demonstrated efferent projections from the NTS to the nucleus paragigantocellularis (PGi) and the central nucleus of the amygdala (CNA) in rat brain. To further characterize the neural circuitry originating from the NTS with postsynaptic targets in the amygdala and medullary autonomic targets, distinct green or red fluorescent latex microspheres were injected into the PGi and the CNA, respectively, of the same rat. Thirty-micron thick tissue sections through the lower brainstem and forebrain were collected. Every fourth section through the NTS region was processed for immunocytochemical detection of tyrosine hydroxylase (TH), a marker of catecholaminergic neurons. Retrogradely labeled neurons from the PGi or CNA were distributed throughout the rostro-caudal segments of the NTS. However, the majority of neurons containing both retrograde tracers were distributed within the caudal third of the NTS. Cell counts revealed that approximately 27% of neurons projecting to the CNA in the NTS sent collateralized projections to the PGi while approximately 16% of neurons projecting to the PGi sent collateralized projections to the CNA. Interestingly, more than half of the PGi and CNA-projecting neurons in the NTS expressed TH immunoreactivity. These data indicate that catecholaminergic neurons in the NTS are poised to simultaneously coordinate activities in limbic and medullary autonomic brain regions.

Caffeine increases paragigantocellularis neuronal firing rate and induces withdrawal signs in morphine-dependent rats

Using single unit recording in nucleus paragigantocellularis neurons located in the rostral ventrolateral medulla, and measuring the precipitated withdrawal syndrome, we investigated whether chronic morphine administration would produce adaptive changes in the adenosine system. Caffeine (50 mg/kg, i.p.) induced withdrawal signs (head shakes, tooth chattering, ejaculation, chewing, and irritability) in morphine-dependent rats 10–18 min after the injection. Only the tooth chattering and diarrhea were expressed following a direct paragigantocellularis injection of caffeine (200 μM, 0.5 μl). The spontaneous activity of paragigantocellularis neurons was significantly decreased by microinjection of both adenosine (10 nM) and an adenosine A 1 receptor-selective agonist, cyclohexyladenosine (200 μM), into the paragigantocellularis nucleus of both control and morphine-dependent rats, but the decrease in firing rate of paragigantocellularis neurons of morphine-dependent rats was greater than that of control ones. There was also a significant enhancement of spontaneous activity of paragigantocellularis neurons 8–15 min after caffeine administration (50 mg/kg, i.p.) and 10–18 min after the injection of an adenosine A 1 receptor-selective antagonist 8-phenyltheophylline (10 mg/kg, i.p.) in both control and morphine-dependent rats. However, the effect of the antagonists was greater in morphine-dependent rats than in control ones. These data suggest that there is an increase in the sensitivity of nucleus paragigantocellularis neurons to adenosine receptor ligands in morphine-dependent rats that may be associated with the ability of caffeine to produce withdrawal signs.

Occurrence of morphine tolerance and dependence in the nucleus paragigantocellularis neurons

The occurrence of morphine tolerance and dependence in the nucleus paragigantocellularis neurons was investigated. The spontaneous activity was recorded from the nucleus paragigantocellularis neurons of urethane-anesthetized rats, using single unit recording. Morphine microinjected (20 mg/ml, 120–200 nl) into the nucleus paragigantocellularis of control rats had both excitatory and inhibitory effects. These effects were reversed by microinjection of naloxone, revealing the possible involvement of μ receptors. Morphine microinjected into morphine-dependent rats failed to change the spontaneous activity of the nucleus paragigantocellularis neurons that accounts for the occurrence of tolerance to morphine in these neurons. Microinjection of naloxone (25 mg/ml, 120–200 nl) in control rats had no effect on the spontaneous firing rate of the nucleus paragigantocellularis neurons but in morphine-dependent rats, either alone or after morphine microinjection, naloxone increased neuronal activity significantly, indicating the occurrence of dependence on morphine in the nucleus paragigantocellularis neurons. These data show that the nucleus paragigantocellularis neurons may play a role in physical dependence on morphine. This conclusion is consistent with the finding, that activation of the nucleus paragigantocellularis by electrical stimulation in morphine-naive rats can elicit behaviors similar to those observed during naloxone-precipitated morphine withdrawal.

Morphine tolerance and dependence in the nucleus paragigantocellularis: single unit recording study in vivo

In this study, a single unit activity was recorded in the nucleus paragigantocellularis (PGi), located in the rostral ventrolateral medulla of anesthetized, morphine-dependent rats. The spontaneous activity of PGi neurons was significantly decreased by administration of morphine (10 mg/kg; i.p.) in sham-operated, control and morphine-dependent rats. However, in PGi neurons of morphine-dependent rats, the firing rate decreased significantly less than those of sham-operated and control ones. There was also significant enhancement of spontaneous activity of PGi neurons for 30 min following administration of naloxone (2 mg/kg; s.c.) in morphine-dependent rats as an opiate withdrawal-induced activation of PGi neurons. The results indicated the occurrence of morphine tolerance and dependence in the PGi and/or elsewhere which appeared in PGi unit activity. The findings are consistent with the hypothesis that during morphine withdrawal, there is an increase in unit activity of the PGi afferents to the nucleus locus coeruleus (LC) or an increased release of excitatory transmitter from their nerve terminals in the LC.

Responsiveness of the nucleus reticularis paragigantocellularis neurons to the formalin as a peripheral noxious stimulus

Responsiveness of the nucleus reticularis paragigantocellularis neurons to the formalin as a peripheral noxious stimulus

Light and electron microscopic evidence for topographic and monosynaptic projections from neurons in the ventral medulla to noradrenergic dendrites in the rat locus coeruleus

Physiological studies have shown that afferents from the nucleus paragigantocellularis (PGi) in the rostral ventral medulla underlie the modulation of locus coeruleus (LC) activity by a variety of stimuli. However, there have been no anatomical demonstrations of a monosynaptic projection from neurons in the PGi to the LC. Thus, biotinylated dextran amine (BDA) was iontophoretically injected into the ventral medulla and single-tissue sections were processed for peroxidase localization of BDA and gold–silver labeling of tyrosine hydroxylase (TH). Discrete microinjections of BDA were placed into either the medial or lateral aspects of the ventral medulla. For medially placed injections, a medio-dorsal pathway to the LC was observed. This trajectory resulted in a predominant innervation of the ventral LC. Lateral injection placements yielded a fiber pathway that coursed more laterally within the medullo-pontine reticular formation and primarily innervated the dorsolateral LC. These light microscopic data suggested that neurons in the PGi use distinct pathways to innervate the LC and are topographically organized within this structure. Electron microscopic analyses of the LC region indicated that axon terminals originating from either subregion were equally likely to contact noradrenergic neurons in the LC. Approximately 57% and 62% of BDA-labeled terminals originating from the medial ( n=150) or lateral ( n=150) aspects of the ventral medulla, respectively, formed heterogeneous synaptic contacts (i.e., inhibitory- and excitatory-type) with dendrites containing TH. It is well known that the PGi is a functionally diverse region that is involved in sensory integration, autonomic regulation and pain modulation. It is also known that LC efferents are spatially organized with respect to their postsynaptic targets. Taken together, our findings that subdivisions of the ventral medulla topographically and monosynaptically innervate the LC suggest that regionally specific PGi neurons target subsets of LC neurons with efferent targets that may possess analogous functional correlates.

The stimulatory effect of clonidine on locus coeruleus neurons of rats with inactivated alpha 2-adrenoceptors: involvement of imidazoline receptors located in the nucleus paragigantocellularis.

Clonidine stimulates locus coeruleus neurons by an alpha 2-adrenoceptor-independent mechanism which probably involves imidazoline receptors. To study this effect, single-unit extracellular recordings in the locus coeruleus were performed in anaesthetised rats after complete, irreversible inactivation of alpha 2-adrenoceptors by the alkylating agent N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) (6 mg/kg i.p.; 6 h before experiments). After this pretreatment, clonidine applied into the locus coeruleus failed to produce any change in the cell firing rate. However, clonidine applied intravenously (320-2560 micrograms/kg), or locally (0.5-2.0 microliters of 0.02 M) into the nucleus paragigantocellularis, a major locus coeruleus afferent, stimulated locus coeruleus neurons (increasing the firing rate by approximately 90%). Electrical lesions of the nucleus paragigantocellularis greatly attenuated the clonidine induced stimulation of locus coeruleus neurons ipsilateral to the lesion when applied intravenously. Blood pressure which was recorded simultaneously with cell recording, remained unaffected after clonidine administration in EEDQ pretreated rats. These results indicate that the clonidine-induced stimulation of locus coeruleus neurons is an indirect effect mediated by imidazoline receptors located on paragigantocellularis neurons projecting to the locus coeruleus.

Robust enkephalin innervation of the locus coeruleus from the rostral medulla

Substantial evidence indicates that the noradrenergic nucleus locus coeruleus (LC) is a key target of endogenous opioid neurons, and an important structure in mediating opiate effects. However, the detailed distribution of opioid fibers and terminals in the LC, and the sources of its opioid innervation are unknown. In the present study, the enkephalin innervation of the LC was investigated in the rat using an antibody directed against the extended enkephalin peptide Met-enkephalin-Arg6-Gly7-Leu8 (ENK), which is derived exclusively from the enkephalin precursor proenkephalin A. An antibody directed against tyrosine hydroxylase (TH), the synthetic enzyme for catecholaminergic neurons, was also applied to the same tissue sections to delineate LC neurons and their dendrites. Enkephalin fibers in the LC were dense and highly varicose. In horizontal sections, ENK-like-immunoreactive (ENK-ir) fibers of considerable length coursed throughout the rostrocaudal orientation of the LC proper, whereas in frontal sections ENK-ir processes appeared punctate, suggesting a rostrocaudal orientation. Dense ENK-ir fibers were also identified in the rostromedial and caudal juxtaependymal pericoerulear regions where extranuclear dendrites of LC neurons are extensive. As previously reported, there were no ENK-ir neurons in the LC nucleus proper, but such cells were present in neighboring structures such as the parabrachial, sphenoid, and Barrington's nuclei as well as in the central gray and in the subcoeruleus area. ENK-ir neurons were also present in nuclei of the rostral medulla reported to be major afferents of the LC, the nucleus prepositus hypoglossi (PrH), and the nucleus paragigantocellularis (PGi). In the dorsomedial medulla, numerous ENK-ir neurons were identified in the medial aspect of the PrH and along the medial longitudinal fasciculus in the perifascicular reticular formation. In the ventrolateral medulla, ENK-ir neurons were distributed in a conical caudorostral column throughout the PGi. Retrograde transport of a WGA-colloidal gold conjugate (WGA-apoHRP-Au) from LC, combined with immunohistochemistry for ENK in the same tissue sections, revealed that LC afferents in the PGi and PrH were interdigitated with ENK-ir neurons. Furthermore, an unexpectedly high incidence of doubly labeled neurons were identified in both PGi and PrH. Overall, 57% and 56% of the LC-projecting neurons in PGi and PrH, respectively, were also immunoreactive for ENK, suggesting that enkephalinergic neurons of PGi and PrH are major afferents to noradrenergic LC neurons.

Adrenergic innervation of the rat nucleus locus coeruleus arises predominantly from the C1 adrenergic cell group in the rostral medulla

Focal iontophoretic injections of the retrograde tracer Fluoro-Gold into the locus coeruleus were combined with immunocytochemistry for phenylethanolamine N-methyltransferase, the final enzyme in the synthesis of epinephrine. Retrograde labeling confirmed recent findings that the major afferents to the locus coeruleus are present in the ventrolateral (nucleus paragigantocellularis) and dorsomedial medulla (nucleus prepositus hypoglossi), areas containing the C1 and C3 adrenergic cell groups, respectively. The Fluoro-Gold label revealed morphologic details of locus coeruleus afferent cells. Labeled neurons in the prepositus hypoglossi region were typically round (10 μm diameter) or ellipsoidal and compressed against the ventricle wall ( 10 × 20μm), while those in the paragigantocellularis were most often multipolar and ellipsoidal or triangular in shape ( 10 × 20−20 × 30μm). Double labeling in the same tissue sections revealed that locus coeruleus afferent neurons are intercalated among phenylethanolamine N-methyltrans-ferase-positive C1 and C3 neurons. Twenty-one per cent of locus coeruleus afferent neurons in paragigantocellularis stained for phenylethanolamine N-methyltransferase while only 4% of locus coeruleus afferents in the prepositus hypoglossi area exhibited phenylethanolamine N-methyltransferase immunoreactivity. In paragigantocellularis, doubly labeled neurons were usually the smaller locus coeruleus afferents, while in the prepositus hypoglossi phenylethanolamine N-methyltransferase labeling was found in all cell types that project to the locus coeruleus. Phenylethanolamine N-methyltransferase-positive fibers from the C1 and C3 cell groups form an adrenergic fiber bundle in the dorsomedial medulla; in the pons, these fibers appear to exit this bundle and innervate the locus coeruleus. Fibers from the neurons of the C3 cell group also appear to ascend on the dorsal surface of the medulla to innervate the locus coeruleus. The phenylethanolamine N-methyltransferase fiber innervation in the locus coeruleus was dense and highly varicose. Phenylethanolamine N-methyltransferase innervation in the dorsal pons was not restricted to the locus coeruleus but was also prominent in neighboring areas such as Barrington's nucleus and the lateral dorsal tegmental nucleus of Gudden.

Two physiologically distinct populations of neurons in the ventrolateral medulla innervate the locus coeruleus

Recent anatomic studies indicate that the nucleus paragigantocellularis (PGi), located in the rostral ventrolateral medulla, strongly innervates the locus coeruleus (LC) while no such input derives from the more caudally located lateral reticular nucleus (LRN). In the present study, focal electrical stimulation of the LC was used to antidromically activate neurons in the ventrolateral medulla. A substantial number of PGi neurons were antidromically driven from the ipsilateral LC, while antidromic activation was virtually absent in LRN. Furthermore, several physiologic properties of antidromically driven cells in PGi define two populations within this group of neurons afferent to LC. These findings provide physiologic confirmation of an anatomically identified input to LC.