The unconditioned fear produced by morphine withdrawal is regulated by mu- and kappa-opioid receptors in the midbrain tectum

We have recently shown that morphine withdrawal sensitizes the neural substrates of fear in the midbrain tectum structures-the dorsal periaqueductal gray (dPAG) and inferior colliculus (IC). In the present study, we investigated the role of mu- and kappa-opioid receptors in the mediation of these effects. Periadolescent rats chronically treated with morphine (10 mg/kg; s.c.) twice daily for 10 days were implanted with an electrode glued to a guide-cannula into the dPAG or the IC. Forty-eight hours after the interruption of this treatment, the effects of intra-dPAG or intra-IC microinjections of [D-Ala(2) N-Me-Phe(4) Gly(5)-ol]-enkephalin (DAMGO; 0.6 and 1 nmol/0.2 mu l) - a selective mu-receptor agonist - or nor-binaltorphimine (BNI; 2.5 and 5 mu g/0.2 mu l) - a selective K-receptor antagonist with tardive action - on the freezing and escape thresholds determined by electrical stimulation of the dPAG and the IC were examined. For both structures, morphine withdrawal produced pro-aversive effects. DAMGO and BNI had antiaversive effects when injected into the dPAG and IC of non-dependent rats. In morphine-withdrawn rats, only BNI continued to promote antiaversive effects in both structures. Whereas DAMGO lost its antiaversive efficacy when injected into the dPAG, only its highest dose promoted antiaversive effects in the IC of morphine-withdrawn rats, suggesting the development of an apparent tolerance. Thus, the enhanced reactivity of the midbrain tectum in morphine-withdrawn periadolescent rats may be due, at least partially, to an impairment of the inhibitory influence of mechanisms mediated by mu-receptors on the neural substrates of fear in this region. (C) 2009 Elsevier B.V. All rights reserved.

Involvement of the midbrain tectum in the unconditioned fear promoted by morphine withdrawal

The midbrain rectum structures, dorsal periaqueductal gray (dPAG) and inferior colliculus (IC), are involved in the organization of fear and anxiety states during the exposure to dangerous stimuli. Since opiate withdrawal is associated with increased anxiety in both humans and animals, this study aimed to investigate the possible sensitization of the neural substrates of fear in the midbrain tectum and its influence on the morphine withdrawal-induced anxiety. For the production of drug withdrawal, rats received morphine injections (10 mg/kg; s.c.) twice daily during 10 days. Forty-eight hours after the interruption of the chronic treatment, independent groups were probed in the elevated plus-maze and open-field tests. Additional groups of animals were implanted with a bipolar electrode into the dPAG OF the IC and submitted to the electrical stimulation of these structures for the determination of the freezing and escape thresholds after 48 h of withdrawal. Our results showed that the morphine withdrawal promoted clear-cut levels of anxiety without the somatic signs of opiate withdrawal. Moreover, morphine-withdrawn rats had an increase in the reactivity to the electrical stimulation of the dPAG and the IC. These findings suggest that the increased anxiety induced by morphine withdrawal is associated with the sensitization of the neural substrates of fear in the dPAG and the IC. So, the present results give support to the hypothesis that withdrawal from chronic treatment with morphine leads to fear states possibly engendered by activation of the dPAG and IC, regardless of the production of somatic symptoms. (C) 2008 Elsevier B.V. All rights reserved.

Involvement of midbrain tectum neurokinin-mediated mechanisms in fear and anxiety

Electrical stimulation of midbrain tectum structures, particularly the dorsal periaqueductal gray (dPAG) and inferior colliculus (IC), produces defensive responses, such as freezing and escape behavior. Freezing also ensues after termination of dPAG stimulation (post-stimulation freezing). These defensive reaction responses are critically mediated by γ-aminobutyric acid and 5-hydroxytryptamine mechanisms in the midbrain tectum. Neurokinins (NKs) also play a role in the mediation of dPAG stimulation-evoked fear, but how NK receptors are involved in the global processing and expression of fear at the level of the midbrain tectum is yet unclear. The present study investigated the role of NK-1 receptors in unconditioned defensive behavior induced by electrical stimulation of the dPAG and IC of male Wistar rats. Spantide (100 pmol/0.2 μL), a selective NK-1 antagonist, injected into these midbrain structures had anti-aversive effects on defensive responses and distress ultrasonic vocalizations induced by stimulation of the dPAG but not of the IC. Moreover, intra-dPAG injections of spantide did not influence post-stimulation freezing or alter exploratory behavior in rats subjected to the elevated plus maze. These results suggest that NK-1 receptors are mainly involved in the mediation of defensive behavior organized in the dPAG. Dorsal periaqueductal gray-evoked post-stimulation freezing was not affected by intra-dPAG injections of spantide, suggesting that NK-1-mediated mechanisms are only involved in the output mechanisms of defensive behavior and not involved in the processing of ascending aversive information from the dPAG.

Glutamatergic neurotransmission mediated by NMDA receptors in the inferior colliculus can modulate haloperidol-induced catalepsy

The inferior colliculus (IC) is primarily involved in the processing of auditory information, but it is distinguished from other auditory nuclei in the brainstem by its connections with structures of the motor system. Functional evidence relating the IC to motor behavior derives from experiments showing that activation of the IC by electrical stimulation or excitatory amino acid microinjection causes freezing, escape-like behavior, and immobility. However, the nature of this immobility is still unclear. The present study examined the influence of excitatory amino acid-mediated mechanisms in the IC on the catalepsy induced by the dopamine receptor blocker haloperidol administered systemically (1 or 0.5 mg/kg) in rats. Haloperidol-induced catalepsy was challenged with prior intracollicular microinjections of glutamate NMDA receptor antagonists, MK-801 (15 or 30 mmol/0.5 mu l) and AP7 (10 or 20 nmol/0.5 mu l), or of the NMDA receptor agonist N-methyl-D-aspartate (NMDA, 20 or 30 nmol/0.5 mu l). The results showed that intracollicular microinjection of MK-801 and AP7 previous to systemic injections of haloperidol significantly attenuated the catalepsy, as indicated by a reduced latency to step down from a horizontal bar. Accordingly, intracollicular microinjection of NMDA increased the latency to step down the bar. These findings suggest that glutamate-mediated mechanisms in the neural circuits at the IC level influence haloperidol-induced catalepsy and participate in the regulation of motor activity. (C) 2010 Published by Elsevier B.V.

Selective involvement of GABAergic mechanisms of the dorsal periaqueductal gray and inferior colliculus on the memory of the contextual fear as assessed by the fear potentiated startle test

The inferior colliculus (IC) together with the dorsal periaqueductal gray (dPAG), the amygdala and the medial hypothalamus make part of the brain aversion system, which has mainly been related to the organization of unconditioned fear. However, the involvement of the IC and dPAG in the conditioned fear is still unclear. It is certain that GABA has a regulatory role on the aversive states generated and elaborated in these midbrain structures. In this study, we evaluated the effects of injections of the GABA-A receptor agonist muscimol (1.0 and 2.0 nmol/0.2 mu L) into the IC or dPAG on the freezing and fear-potentiated startle (FPS) responses of rats submitted to a context fear conditioning. Intra-IC injections of muscimol did not cause any significant effect on the FPS or conditioned freezing but enhanced the startle reflex in non-conditioned animals. In contrast, intra-dPAG injections of muscimol caused significant reduction in FPS and conditioned freezing without changing the startle reflex in non-conditioned animals. Thus, intra-dPAG injections of muscimol produced the expected inhibitory effects on the anxiety-related responses, the FPS and the freezing whereas these injections into the IC produced quite opposite effects suggesting that descending inhibitory pathways from the IC, probably mediated by GABA-A mechanisms, exert a regulatory role on the lower brainstem circuits responsible for the startle reflex. (C) 2008 Elsevier Inc. All rights reserved.

Anxiety-like symptoms induced by morphine withdrawal may be due to the sensitization of the dorsal periaqueductal grey

Withdrawal from morphine leads to the appearance of extreme anxiety accompanied of several physical disturbances, most of them linked to the activation of brainstem regions such as the locus coeruleus, ventral tegmental area, hypothalamic nuclei and periaqueductal grey (PAG). As anxiety remains one of the main components of morphine withdrawal the present study aimed to evaluating the influence of the dorsal aspects of the PAG on the production of this state, since this structure is well-known to be involved in defensive behaviour elicited by anxiety-evoking stimuli. Different groups of animals were submitted to 10 days of i.p. morphine injections, challenged 2 h after with an i.p. injection of naloxone (0.1 mg/kg), and submitted to the plus-maze, open-field and light-dark transition tests. The effects of morphine withdrawal on anxiety-induced Fos immunolabelling were evaluated in four animals that passed by the light-dark transition test randomly chosen for Fos-protein analysis. Besides the PAG, Fos neural expression was conducted in other brain regions involved in the expression of anxiety-related behaviours. Our results showed that morphine withdrawn rats presented enhanced anxiety accompanied of few somatic symptoms. Increased Fos immunolabelling was noted in brain regions well-known to modulate these states as the prelimbic cortex, nucleus accumbens, amygdala and paraventricular hypothalamus. Increased Fos labelling was also observed in the ventral and dorsal aspects of the PAG, a region involved in anxiety-related processes suggesting that this region could be a common neural substrate enlisted during anxiety evoked by dangerous stimuli as well as those elicited by opiate withdrawal. (c) 2008 Elsevier Inc. All rights reserved,

Regulation of conditioned and unconditioned fear in rats by 5-HT1A receptors in the dorsal periaqueductal gray

Studies on the involvement of 5-HT1-mediated mechanisms in the dorsal periaqueductal gray (dPAG) of animals with past stressful experiences have not been conducted so far. We investigated the role of 5-HT1 receptors in the dPAG of rats previously submitted to contextual fear conditioning. Defensive behaviors induced by activation of the dPAG were assessed by measuring the lowest electric current applied to this structure (threshold) able to produce freezing and escape responses during testing sessions of contextual fear conditioning, in which animals were placed in a context previously paired to footshocks. The 5-HT1A function of the dPAG was evaluated by local injections of 8-OH-DPAT (4 and 8 nmol/0.2 mu L) and WAY-100635 (10 nmol/0.2 mu L), selective agonist and antagonist of 5-HT1A receptors, respectively. In accordance with previous studies, 8-OH-DPAT increased aversive thresholds (antiaversive effects) but injections of WAY 100635 into the dPAG did not produce significant effects on the aversive thresholds in naive rats. However, the aversive thresholds of animals exhibiting contextual fear remained unchanged with both treatments. Moreover, 8-OH-DPAT and WAY 100635 did not change the dPAG post-stimulation freezing. The present results suggest that the stressful experience of being fear conditioned has an effect on the role of the 5-HT1A receptors in mediating unconditioned fear. Also, the reduction in the regulation of the defensive behaviors by 5-HT1A-mediated mechanisms in the dPAG of these animals may underlie the stress precipitated psychopathology associated with the neural substrates of aversion of the dPAG. (c) 2007 Elsevier Inc. All rights reserved.

Conditioned fear is modulated by D(2) receptor pathway connecting the ventral tegmental area and basolateral amygdala

Excitation of the mesocorticolimbic pathway, originating from dopaminergic neurons in the ventral tegmental area (VTA), may be important for the development of exaggerated fear responding. Among the forebrain regions innervated by this pathway, the amygdala is an essential component of the neural circuitry of conditioned fear. The functional role of the dopaminergic pathway connecting the VIA to the basolateral amygdala (BLA) in fear and anxiety has received little attention. In vivo microdialysis was performed to measure dopamine levels in the BLA of Wistar rats that received the dopamine D(2) agonist quinpirole (1 mu g/0.2 mu l) into the VTA and were subjected to a fear conditioning test using a light as the conditioned stimulus (CS). The effects of intra-BLA injections of the D(1) antagonist SCH 23390 (1 and 2 mu g/0.2 mu l) and D(2) antagonist sulpiride (1 and 2 mu g/0.2 mu l) on fear-potentiated startle (FPS) to a light-CS were also assessed. Locomotor performance was evaluated by use of open-field and rotarod tests. Freezing and increased dopamine levels in the BLA in response to the CS were both inhibited by intra-VTA quinpirole. Whereas intra-BLA SCH 23390 did not affect FPS, intra-BLA sulpiride (2 mu g) inhibited FPS. Sulpiride`s ability to decrease FPS cannot be attributed to nonspecific effects because this drug did not affect motor performance. These findings indicate that the dopamine D(2) receptor pathway connecting the ventral tegmental area and the basolateral amygdala modulates fear and anxiety and may be a novel pharmacological target for the treatment of anxiety. (C) 2010 Elsevier Inc. All rights reserved.

GABAergic REGULATION OF AUDITORY SENSORY GATING IN LOW- AND HIGH-ANXIETY RATS SUBMITTED TO A FEAR CONDITIONING PROCEDURE

The inferior colliculus (IC) is primarily involved in the processing of acoustic stimuli, being in a position to send auditory information to motor centers that participate in behaviors such as prey catching and predators` avoidance The role of the central nucleus of the IC (CIC) on fear and anxiety has been suggested on the basis that rats are able to engage in tasks to decrease the aversiveness of CIC stimulation, increased Fos immunolabeling during diverse aversive states and increased CIC auditory evoked potentials (AEP) induced by conditioned fear stimuli Additionally it was shown that brainstem AEP, represented by wave V, for which the main generator is the IC, is increased during experimentally induced anxiety Rats segregated according to their low or high emotional reactivity have been used as an important tool in the study of fear and anxiety The IC contains a high density of GABA receptors Since the efficacy of an anxiolytic compound is a function of the animal`s anxiety level, it is possible that GABA-benzodiazepine (Bzp) agents affect LA and HA animals differently In this study we investigated the GABA-Bzp influence on the modulation of AEP in rats with low (LA) or high-anxiety (HA) levels, as assessed by the elevated plus maze test (EPM) GABA-Bzp modulation on the unconditioned AEP response was analyzed by using intra CIC injections (0 2 mu l) of the GABA-Bzp agonists muscimol (121 ng) and diazepam (30 mu g) or the GABA inhibitors bicuculline (10 ng) and semicarbazide (7 mu g) In a second experiment, we evaluate the effects of contextual aversive conditioning on AEP using foot shocks as unconditioned stimuli On the unconditioned fear paradigm GABA inhibition in creased AEP in LA rats and decreases this measure in HA counterparts Muscimol was effective in reducing AEP in both LA and HA rats Contextual fear stimuli increased the magnitude of AEP In spite of no effect obtained with diazepam in LA rats the drug inhibited AEP in HA animals The specificity of the regulatory mechanisms mediated by GABA Bzp for the ascending neurocircuits responsible for the acquisition of aversive information in LA and HA animals shed light on the processing of sensory information underlying the generation of defensive reactions (C) 2010 IBRO Published by Elsevier Ltd All rights reserved

Intrinsic changes in developing retinal neurons result in regenerative failure of their axons.

The failure of mature mammalian central nervous system axons to regenerate after transection is usually attributed to influences of the extraneuronal milieu. Using explant cocultures of retina and midbrain tectum from hamsters, we have found evidence that these influences account for failure of regrowth of only a small minority of retinal axons. For most of the axons, there is a programmed loss of ability to elongate in the central nervous system. We show that there is a precipitous decline in the ability of retinal axons to reinnervate tectal targets when the retina is derived from pups on or after postnatal day 2, even when the target is embryonic. By contrast, embryonic retinal axons can regrow into tectum of any age, overcoming growth-inhibiting influences of glial factors.

Evolutionary Changes in the Complexity of the Tectum of Nontetrapods: A Cladistic Approach

Background: The tectum is a structure localized in the roof of the midbrain in vertebrates, and is taken to be highly conserved in evolution. The present article assessed three hypotheses concerning the evolution of lamination and citoarchitecture of the tectum of nontetrapod animals: 1) There is a significant degree of phylogenetic inertia in both traits studied (number of cellular layers and number of cell classes in tectum); 2) Both traits are positively correlated accross evolution after correction for phylogeny; and 3) Different developmental pathways should generate different patterns of lamination and cytoarchitecture.Methodology/Principal Findings: The hypotheses were tested using analytical-computational tools for phylogenetic hypothesis testing. Both traits presented a considerably large phylogenetic signal and were positively associated. However, no difference was found between two clades classified as per the general developmental pathways of their brains.Conclusions/Significance: The evidence amassed points to more variation in the tectum than would be expected by phylogeny in three species from the taxa analysed; this variation is not better explained by differences in the main course of development, as would be predicted by the developmental clade hypothesis. Those findings shed new light on the evolution of an functionally important structure in nontetrapods, the most basal radiations of vertebrates.

DCC plays a role in navigation of forebrain axons across the ventral midbrain commissure in embryonic Xenopus

In the developing vertebrate brain, growing axons establish a scaffold of axon tracts connected across the midline via commissures. We have previously identified a population of telencephalic neurons that express NOC-2, a novel glycoform of the neural cell adhesion molecule N-CAM that is involved in axon guidance in the forebrain. These axons arise from the presumptive telencephalic nucleus, course caudally along the principal longitudinal tract of the forebrain, cross the ventral midline in the midbrain, and then project to the contralateral side of the brain. In the present study we have investigated mechanisms controlling the growth of these axons across the ventral midline of the midbrain. The axon guidance receptor DCC is expressed by the NOC-2 population of axons both within the longitudinal tract and within the ventral midbrain commissure. Disruption of DCC-dependent interactions, both in vitro and in vivo, inhibited the NOC-2 axons from crossing the ventral midbrain. Instead, these axons grew along aberrant trajectories away from the midline, suggesting that DCC-dependent interactions are important for overcoming inhibitory mechanisms within the midbrain of the embryonic vertebrate brain. Thus, coordinated responsiveness of forebrain axons to both chemostimulatory and chemorepulsive cues appears to determine whether they cross the ventral midline in the midbrain, (C) 2000 Academic Press.

Adrenalectomy counteracts the local modulation of astroglial fibroblast growth factor system without interfering with the pattern of 6-OHDA-induced dopamine degeneration in regions of the ventral midbrain

The present study investigated the effects of bilateral adrenalectomy (ADX) on the synthesis of basic fibroblast growth factor (bFGF, FGF-2) mRNA and on the expression of its FGF receptor subtype-2 (FGFR2) mRNA after a 6-hydroxydopamine (6-OHDA)-induced lesion of nigrostriatal dopamine system. In previous papers we have demonstrated that corticosterone increases FGF-2 immunoreactivity mainly in the astrocytes of the substantia nigra [Chadi, G., Rosen, L., Cintra, A., Tinner, B., Zoli, M., Pettersson, R.F., Fuxe, K., 1993b. Corticosterone increases FGF-2 (bFGF) immunoreactivity in the substantia nigra of the rat. Neuroreport 4, 783-786.] and that 6-OHDA injected in the ventral midbrain upregulates FGF-2 synthesis in reactive astrocytes in the ascending dopamine pathways [Chadi, G., Cao, Y., Pettersson, R.F., Fuxe, K., 1994. Temporal and spatial increase of astroglial basic fibroblast growth factor synthesis after 6-hydroxydopamine-induced degeneration of the nigrostriatal dopamine neurons. Neuroscience 61, 891-910.]. Rats were adrenalectomized and received a 6-OHDA stereotaxical injection in the ventral midbrain 2 days later. Seven days after the dopamine lesion, Western blot analysis showed a decreased level of tyrosine hydroxylase in the lesioned side of the midbrain, an event that was not altered by ADX or corticosterone replacement. Moreover, the degeneration of nigral dopamine neurons, which was confirmed by the disappearance of acidic FGF (FGF-1) mRNA and the decrement of tyrosine hydroxylase mRNA labeled nigral neurons, was not altered by ADX. The FGF-2 protein (23 kDa isoform but not 21 kDa fraction) levels increased in the lesioned side of the ventral midbrain. This elevation was counteracted by ADX, an effect that was fully reversed by corticosterone replacement. In situ hybridization revealed that ADX counteracted the elevated FGF-2 mRNA levels in putative glial cells of the ipsilateral pars compacta of the substantia nigra and in the ventral tegmental area. The ADX also counteracted the increased density and intensity of the astroglial FGF-2 immunoreactive profiles within the lesioned pars compacta of the substantia nigra and the ventral tegmental area as determined by stereology. The stereotaxical mechanical needle insertion triggered the expression of FGFR2 mRNA in putative glial cells, spreading to the entire ipsilateral ventral midbrain from the region of needle track, an occurrence that was partially reversed by ADX. In conclusion, bilateral ADX counteracted the increased astroglial FGF-2 synthesis in the dopamine regions of the ventral midbrain following a 6-OHDA-induced local lesion and interfered with FGF receptor regulation around injury. These findings give further evidence that adrenocortical hormones may regulate the astroglial FGF-2-mediated trophic mechanisms and wound repair events in the lesioned central nervous system. (c) 2007 Elsevier B.V. All rights reserved.

Task-dependent modulation of reward and novelty processing in human ventral striatum and midbrain

Magdeburg, Univ., Fak. für Naturwiss., Diss., 2008

Extent of Hypoattenuation On CT Angiography Source Images In Basilar Artery Occlusion: Prognostic Accuracy Of Posterior Circulation ASPECTS And The Pons-midbrain Score In The Basilar Artery International Cooperation Study

Background: The posterior circulation Acute Stroke Prognosis Early CT Score (pc-ASPECTS) and the combined Pons-midbrain score quantify the extent of early ischemic changes in the posterior circulation. We compared the prognostic accuracy of both scores if applied to CT angiography (CTA) source images (CTA-SI) of patients in the Basilar Artery International Cooperation Study (BASICS).Methods: BASICS was a prospective, observational, multi-centre, registry of consecutive patients who presented with acute symptomatic basilar artery occlusion (BAO). Functional outcome was assessed at 1 month. We applied pc-ASPECTS and the combined Pons-midbrain score to CTA-SI by 3-reader-consensus. Readers were blinded to clinical data. We performed multivariable logistic regression analysis, adjusting for thrombolysis, baseline NIHSS score and age, and used the output to derive ROC curves to compare the ability of both scores to discriminate patients with favourable (modified Rankin Scale [mRS] scores 0-3) from patients with unfavourable (mRS scores 4-6) functional outcome.Results: We reviewed CTAs of 158 patients (64% men, mean age 65 _ 15 years, median NIHSS score 25 [0-38], median GCS score 7 [3-15], median onset-to-CTA time 234 minutes [11-7380]). At 1 month, 40 (25%) patients had a favourable outcome, 49 (31%) had an unfavourable outcome (mRS score 4-5) and 69 (44%) were deceased. Both techniques of assessing CTA-SI hypoattenuation in the posterior circulation showed equally good discriminative value in predicting final outcome (C-statistics; area under ROC curve 0.74 versus 0.75, respectively; p_0.37). Pc-ASPECTS dichotomized at _6 versus _6 was an independent predictor of favourable functional outcome (RR _ 2.2; CI95 1.1-4.7; p _ 0.034).Conclusion: Compared to the combined Pons-midbrain score, the pc-ASPECTS score has similar prognostic accuracy to identify patients with a favourable functional outcome in BASICS. Dichotomized pc-ASPECTS (_6 versus _6) is an independent predictor of favourable functional outcome in this population. Author Disclosures: V. Puetz: None. A. Khomenko: None. M.D. Hill: None. I. Dzialowski: None. P. Michel: None. C. Weimar: None. C.A.C. Wijman: None. H. Mattle: None. K. Muir: None. T. Pfefferkorn: None. D. Tanne: None. S. Engelter: None. K. Szabo: None. A. Algra: None. A.M. Demchuk: None. W.J. Schonewille: None.