Orexinergic innervation of the extended amygdala and basal ganglia in the rat.

The orexinergic system interacts with several functional states of emotions, stress, hunger, wakefulness and behavioral arousal through four pathways originating in the lateral hypothalamus (LH). Hundreds of orexinergic efferents have been described by tracing studies and direct immunohistochemistry of orexin in the forebrain, olfactory regions, hippocampus, amygdala, septum, basal ganglia, thalamus, hypothalamus, brain stem and spinal cord. Most of these tracing studies investigated the whole orexinergic projection to all regions of the intracranial part of the CNS. To identify the orexinergic efferents at the subnuclear level of resolution, we focussed on the orexinergic target in the amygdala, which is substantially involved in the LH output and contributes mostly to the functional outcome of the orexinergic system and the basal ganglia. Immunohistochemical identification of axonal orexin A and orexin B in male adult rats has been performed on serial sections. In the extended amygdala many new orexinergic targets were found in the anterior amygdaloid area (dense), anterior cortical nucleus (moderate), amygdalostriatal transition region (moderate), basolateral regions (moderate), basomedial nucleus (moderate), several bed nucleus of the stria terminals regions (few to dense), central amygdaloid subdivisions (dense), posteromedial cortical nucleus (moderate) and medial amygdaloid subnuclei (dense). Furthermore, the entopeduncular nucleus has been newly identified as another target for orexinergic fibers with a high density. These results suggest that subdivisions and subnuclei of the extended amygdala are specific targets of the orexinergic system.

Amygdalotrigeminal projection in the rat: An anterograde tracing study.

Previous neurophysiological studies have demonstrated that the amygdala has a direct influence upon trigeminal motoneuron activity. The existence of a direct amygdalotrigeminal pathway in rats was proved by anterograde tracing with the neuroanatomical tracer, biotinylated dextran amine (BDA). After ipsilateral BDA application to the central nucleus of the amygdala (AmCe), widespread ipsilateral projections emerging from its medial subnucleus were traced to the trigeminal brainstem nuclear complex, including the principal sensory (Pr5) and mesencephalic trigeminal nucleus (Me5), and their premotoneurons and interneurons, located in the supratrigeminal, intertrigeminal and peritrigeminal nuclei. Sparse BDA-labeled axons and their terminals were also distributed in the contralateral Pr5, interpolar and caudal subnuclei of the spinal trigeminal nucleus. The central lateral amygdaloid nucleus gives rise to a light ipsilateral projection to the pontine part of the Me5. The present data indicate that AmCe sends massive efferents to the trigeminal nuclei in the brainstem, wherein its medial subnucleus sends the major input to them. The medial amygdaloid nucleus sparsely innervates Me5 neurons, specifically those located in its mesencephalic portion, while basomedial and basolateral efferents do not target the trigeminal nuclear complex. These results suggest that the amygdaloid input may modulate the activity of trigeminal sensory and motor neurons and, thus, the amygdala is possibly involved in the control of masticatory behavior.

Activation-dependent descending reflex evacuation motority of anal canal in rat model.

The evacuative motor responses of the anal canal and recto-anal reflexes during defecation were studied in an isolated rat recto-anal model preparation using (i) partitioned organ bath, (ii) electrical stimulation, (iii) balloon distension and (iv) morphological techniques. Electrical field stimulation applied to the anal canal or to the distal part of the rectum elicited tetrodotoxin (10(-7) M)-sensitive frequency-dependent local or descending contractions of the anal canal and the local responses were bigger in amplitude (14.9 ± 1.35 mN) than the descending contractions (5.3 ± 0.7 mN at frequency of 5 Hz, p < 0.05). The balloon-induced distension of the distal rectum evoked descending responses of the anal canal consisting of a short contraction (1.50 ± 0.18 mN) followed by deep relaxation (3.12 ± 0.34 mN). In the presence of atropine (3 x 10(-7) M) the electrically-elicited (5 Hz) local or descending contractions of the anal canal were suppressed and a relaxation revealed. The initial contraction component of the distension-induced response was decreased while the relaxation was not changed. During atropine treatment, spantide (10(-7) M) lowered even more the contractile component of the anal canal response. NG-nitro-L-arginine (5 x 10(-4) M) enhanced the contraction, prevented the atropine-dependent relaxation of the electrically-elicited response and inhibited the distension-induced relaxation. L-Arginine (5 x 10(-4) M) suppressed the contraction and extended the relaxation. ChAT-, substance P- and NADPH-diaphorase-positive perikarya and nerve fibers were observed in myenteric ganglia of the anal canal. The results suggest activation-dependent descending reflex motority of the anal canal involving electrical stimulation-displayed cholinergic and tachykininergic and distension manifested nitrergic neuro-muscular communications.

Diurnal rhythms of spontaneous recurrent seizures and behavioral alterations of Wistar and spontaneously hypertensive rats in the kainate model of epilepsy.

Attention deficit hyperactivity disorder (ADHD) can coexist with epilepsy. Spontaneously hypertensive rats (SHRs) are considered to model ADHD with overactivity, impulsiveness, deficient sustained attention, and alterations in circadian autonomic profiles. The present study explored spontaneous recurrent seizures (SRSs) and behavioral diurnal activity rhythms in normotensive Wistar rats and SHRs in the kainate model of epilepsy. Rats were video monitored (24 h/3 months) to detect SRSs. SHRs manifested a lower seizure frequency during the light phase in the 8th and 10th weeks and a lower frequency of SRSs during the night phase accompanied by attenuated responses in hyperexcitability tests. Both epileptic strains were hyperactive, with lower anxiety levels, and their diurnal rhythms were abolished. Epileptic Wistar rats and SHRs exhibited less exploration during the dark phase. This study suggests that SHRs may be useful in modeling some aspects (particularly hypertension-related diurnal rhythm disturbance) of behavior associated with epilepsy.

Locality-dependent descending reflex motor activity in the anal canal--cholinergic and nitrergic contributions in the rat model.

AIM: Since the distal part of the intestine is targeted by a wide range of pathogens, the motility of the recto-anal region has been the object of many experimental and clinical observations. In this study, we investigated descending motor responses in the anal canal as a measure of the activation of autonomic reflex pathways underlying evacuatory recto-anal activity. METHODS: The partitioned organ bath method was used to register motor responses of the anal canal as induced by balloon distension of the rectum in isolated rat recto-anal preparations. RESULTS: Distension-induced descending responses of the anal canal comprised contractions (with distension at a distance of 15 mm), initial contractions and secondary relaxations (at 10 mm) and short contractions followed by deep relaxations (at 3-5 mm). Decreasing the distance between the distension stimulus and the anal canal resulted in a decreased contraction response and increased relaxation. Tetrodotoxin (0.1 micromol/L) inhibited these responses. Atropine (0.3 micromol/L) decreased contraction and did not change the relaxation response. N(G)-nitro-L-arginine (0.5 mmol/L) enhanced contraction in both the absence and presence of atropine. L-arginine (0.5 mmol/L) inhibited contraction and extended relaxation in atropine-pretreated preparations. The actions of N(G)-nitro-L-arginine and L-arginine were more pronounced in the aboral direction. ChAT-positive nerve fibers were observed in myenteric ganglia of the rectum and the anal canal. The density of NADPH-diaphorase-positive neurons was higher in the anal canal region. CONCLUSION: Our results suggest that locality-dependent activation of the descending reflex neuromuscular communications underlie evacuatory activity in the recto-anal region. This activation response involves long excitatory cholinergic and non-cholinergic pathways along the rectum and short inhibitory nitrergic pathways located predominantly in the anal canal region.

Efferent projections of the anterior and posterodorsal regions of the medial nucleus of the amygdala in the mouse.

The efferent projections of the anterior and posterodorsal part of the medial nucleus (MePD) in the mouse were studied by means of anterograde axonal tracing using biotinylated dextran amine. The MePD axons ran mainly via the stria terminalis and to a lesser extent via the ventral amygdalofugal pathway. The projections to the forebrain were broadly distributed and varied from very strong to scant. The most significant connections were destined to the bed nucleus of the stria terminalis in which all parts of the medial division were innervated by MePD neurons. Moderate projections reached the limbic striatum (nucleus accumbens), olfactory tubercle and the lateral septal nucleus. The substantia innominata was also innervated by the MePD, and especially the projection to its ventral portion was substantial. The profuse innervation of the medial preoptic nucleus and medial preoptic area indicated significant involvement of the MePD in sexual behavior. Many hypothalamic nuclei were innervated but to a different extent. The very strong innervation of the ventral premammillary nucleus further indicated the involvement of the MePD in the neuronal circuitry for sexual behavior. Substantial projections also reached the anterior hypothalamus and tuber cinereum, while the connections to the lateral hypothalamus were widespread but showed moderate density. MePD strongly innervated the ventrolateral part of the ventromedial hypothalamic nucleus and moderately its remaining parts. The neurosecretory hypothalamic nuclei and the arcuate nucleus contained only a few MePD terminals. The thalamic innervation was very scant and reached the lateral habenular nucleus and the nuclei of the midline. The mesencephalic connections were moderate to sparse and projected to the mesolimbic dopaminergic groups in the ventral tegmental area, the pars lateralis and the dorsal tier of the substantia nigra pars compacta, the periaqueductal gray and the dorsal raphe nucleus. The present results principally resembled data known in other rodent species; however, the efferents of the MePD often differed in extent and/or topical distribution.

Ascending and descending reflex motor activity of recto-anal region-cholinergic and nitrergic implications in a rat model.

The implications of cholinergic and nitrergic transmissions in ascending and descending reflex motor pathways of recto-anal region in rat model were evaluated using: (i) electrical stimulation; (ii) triple organ bath; and (iii) morphological techniques. Electrical stimulation to anal canal induced simultaneous ascending contractile responses of longitudinal and circular muscles of proximal rectum, local contraction of anal canal or contraction followed by relaxation of internal anal sphincter when external sphincter was dissected off. The stimulation of proximal rectum elicited local contractions of both rectal layers and descending contractions of internal sphincter or anal canal. Tetrodotoxin (0.1 microM) prevented the electrically elicited events. The ascending excitatory responses and the local and ascending contractions of longitudinal muscle were more pronounced than those of circular muscle suggesting dominant role of ascending reflex pathways and of longitudinal muscle in rectal motor activity. Choline acetyltransferase (ChAT)-containing fibres and nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase-positive neurons were observed in myenteric ganglia of rectum and anal canal. NG-nitro-l-arginine (0.5mM) increased the contractile ascending and descending responses. During atropine (0.3 microM) treatment the ascending and descending contractions were suppressed but not abolished and a relaxation revealed in ascending response of circular muscle and in descending responses of internal anal sphincter and anal canal. The relaxation was decreased by NG-nitro-l-arginine and increased by l-arginine (0.5mM). The results suggest that cholinergic excitatory ascending and descending pathways and nitric oxide-dependent inhibitory ascending neurotransmission(s) to rectal circular muscle and inhibitory descending to internal anal sphincter and anal canal are involved in reflex circuitry controlling motor activity of recto-anal region.

Efferent connections of the parabigeminal nucleus to the amygdala and the superior colliculus in the rat: a double-labeling fluorescent retrograde tracing study.

The parabigeminal nucleus (Pbg) is a subcortical visual center that besides reciprocal connections with the superior colliculus (SC), also projects to the amygdala (Am). The Pbg-Am connection is part of a multineuronal pathway that conveys extrageniculostriate inputs of the retina to the Am, and it rapidly responds to the sources of threat before conscious detection. The present study demonstrates that Pbg projects bilaterally to Am and SC. The ipsilateral projections arise from separate cell populations, whilst the contralaterally projecting Pbg neurons emit branching axons that simultaneously innervate Am and SC.

Glutamate stimulation of acetylcholine release from myenteric plexus is mediated by endogenous nitric oxide.

Glutamate was found to be an excitatory neurotransmitter in the enteric nervous system. Although several lines of evidence indicate a role of glutamate in the regulation of gut motility and secretion the physiological significance of glutamatergic transmission is not clear yet. We studied the effect of glutamate on [3H]acetylcholine release and nicotinamide adenine dinucleotide phosphate-diaphorase staining in longitudinal muscle strips with attached myenteric plexus of guinea pig ileum. L-glutamate (100 microM) significantly enhanced both the evoked [3H]acetylcholine release and the optical density of nicotinamide adenine dinucleotide phosphate-diaphorase positive neurones, i.e. the intensity of staining. The non-competitive N-methyl-D-aspartate receptor antagonist MK-801 (3 microM) abolished the stimulatory effect of L-glutamate on acetylcholine efflux. Similarly, the nitric oxide synthase inhibitor N(omega)-nitro-L-arginine (100 microM) significantly reduced the effect of L-glutamate on [3H]acetylcholine release and nicotinamide adenine dinucleotide phosphate-diaphorase staining. Our data suggest that endogenous nitric oxide seems to mediate the stimulatory effect of glutamate on acetylcholine release from guinea pig myenteric neurons.

Heme oxygenase-2 products activate IKCa: role of CO and iron in guinea pig portal vein smooth muscle cells.

Hemin (10 microM) and carbon monoxide (CO) increased iberiotoxin-blockable IKCa in portal vein smooth muscle cells. CO-induced IKCa activation was abolished by 10 microM ODQ, 10 microM cyclopiazonic acid and 1 microM KT5823. The hemin-induced effect on IKCa was abolished by pretreatment with Sn-protoporphyrin IX, a heme oxygenase inhibitor and Fe2+ chelator but was insensitive to inhibitors of soluble guanylate cyclase (GC) and cGMP-dependent protein kinase (PKG). There was no effect of hemin on IKCa in the presence of 3 microM dithiotreitol into the bath or 3 mM glutathione into the pipette solution. Superoxide dismutase (1000 U/ml) or catalase (3000 U/ml) added into the pipette solution also abolished the effect of hemin on IKCa in this tissue. Additionally, 10 microM hemin could not influence IKCa in Ca2+-free external solution or in the presence of 30 microM SKF 95356. It was concluded that CO increases IKCa via its "conventional" signaling pathway, which involves soluble GC and PKG activation and subsequent stimulation of sarcoplasmic reticulum Ca2+ pump activity resulting in Ca2+-dependent activation of IKCa due to the accumulation of Ca2+ into the space near the plasma membrane. On the other hand, internally produced CO could not yield the same IKCa increase, while Fe2+ derived from heme oxygenase 2-dependent degradation of hemin in portal vein smooth muscle cells gives rise to reactive oxygen species namely hydroxyl and superoxide radicals. Both radicals are responsible for the SKF 95356-sensitive non-selective cation channel activation, the Ca2+ influx and the subsequent increase of Ca2+ concentration near the plasma membrane that augments the KCa channel activity.