Brain metabolic changes and clinical response to superolateral medial forebrain bundle deep brain stimulation for treatment-resistant depression.

Deep brain stimulation (DBS) to the superolateral branch of the medial forebrain bundle is an efficacious therapy for treatment-resistant depression, providing rapid antidepressant effects. In this study, we use 18F-fluorodeoxyglucose-positron emission tomography (PET) to identify brain metabolic changes over 12 months post-DBS implantation in ten of our patients, compared to baseline. The primary outcome measure was a 50% reduction in Montgomery-Åsberg Depression Rating Scale (MADRS) score, which was interpreted as a response. Deterministic fiber tracking was used to individually map the target area; probabilistic tractography was used to identify modulated fiber tracts modeled using the cathodal contacts. Eight of the ten patients included in this study were responders. PET imaging revealed significant decreases in bilateral caudate, mediodorsal thalamus, and dorsal anterior cingulate cortex metabolism that was evident at 6 months and continued to 12 months post surgery. At 12 months post-surgery, significant left ventral prefrontal cortical metabolic decreases were also observed. Right caudate metabolic decrease at 12 months was significantly correlated with mean MADRS reduction. Probabilistic tractography modeling revealed that such metabolic changes lay along cortico-limbic nodes structurally connected to the DBS target site. Such observed metabolic changes following DBS correlated with clinical response provide insights into how future studies can elaborate such data to create biomarkers to predict response, the development of which likely will require multimodal imaging analysis.

Machine learning-aided personalized DTI tractographic planning for deep brain stimulation of the superolateral medial forebrain bundle using HAMLET.

BACKGROUND: Growing interest exists for superolateral medial forebrain bundle (slMFB) deep brain stimulation (DBS) in psychiatric disorders. The surgical approach warrants tractographic rendition. Commercial stereotactic planning systems use deterministic tractography which suffers from inherent limitations, is dependent on manual interaction (ROI definition), and has to be regarded as subjective. We aimed to develop an objective but patient-specific tracking of the slMFB which at the same time allows the use of a commercial surgical planning system in the context of deep brain stimulation. METHODS: The HAMLET (Hierarchical Harmonic Filters for Learning Tracts from Diffusion MRI) machine learning approach was introduced into the standardized workflow of slMFB DBS tractographic planning on the basis of patient-specific dMRI. Rendition of the slMFB with HAMLET serves as an objective comparison for the refinement of the deterministic tracking procedure. Our application focuses on the tractographic planning of DBS (N = 8) for major depression and OCD. RESULTS: Previous results have shown that only fibers belonging to the ventral tegmental area to prefrontal/orbitofrontal axis should be targeted. With the proposed technique, the deterministic tracking approach, that serves as the surgical planning data, can be refined, over-sprouting fibers are eliminated, bundle thickness is reduced in the target region, and thereby probably a more accurate targeting is facilitated. The HAMLET-driven method is meant to achieve a more objective surgical fiber display of the slMFB with deterministic tractography. CONCLUSIONS: The approach allows overlying the results of patient-specific planning from two different approaches (manual deterministic and machine learning HAMLET). HAMLET shows the slMFB as a volume and thus serves as an objective tracking corridor. It helps to refine results from deterministic tracking in the surgical workspace without interfering with any part of the standard software solution. We have now included this workflow in our daily clinical experimental work on slMFB DBS for psychiatric indications.

Tractography-assisted deep brain stimulation of the superolateral branch of the medial forebrain bundle (slMFB DBS) in major depression.

Background: Deep brain stimulation (DBS) of the superolateral branch of the medial forebrain bundle (slMFB) emerges as a - yet experimental - treatment for major depressive disorder (MDD) and other treatment refractory psychiatric diseases. First experiences have been reported from two open label pilot trials in major depression (MDD) and long-term effectiveness for MDD (50 months) has been reported. Objective: To give a detailed description of the surgical technique for DBS of the superolateral branch of the medial forebrain bundle (slMFB) in MDD. Methods: Surgical experience from bilateral implantation procedures in n = 24 patients with MDD is reported. The detailed procedure of tractography-assisted targeting together with detailed electrophysiology in 144 trajectories in the target region (recording and stimulation) is described. Achieved electrode positions were evaluated based on postoperative helical CT and fused to preoperative high resolution anatomical magnetic resonance imaging (MRI; Philips Medical Systems, Best, Netherlands), including the pre-operative diffusion tensor imaging (DTI) tractographic information (StealthViz DTI, Medtronic, USA; Framelink 5.0, Medtronic, USA). Midcommissural point (MCP) coordinates of effective contact (EC) location, together with angles of entry into the target region were evaluated. To investigate incidental stimulation of surrounding nuclei (subthalamic nucleus, STN; substantia nigra, SNr; and red nucleus, RN) as a possible mechanism, a therapeutic triangle (TT) was defined, located between these structures (based on MRI criteria in T2) and evaluated with respect to EC locations. Results: Bilateral slMFB DBS was performed in all patients. We identified an electrophysiological environment (defined by autonomic reaction, passive microelectrode recording, acute effects and oculomotor effects) that helps to identify the proper target site on the operation table. Postoperative MCP-evaluation of effective contacts (EC) shows a significant variability with respect to localization. Evaluation of the TT shows that responders will typically have their active contacts inside the triangle and that surrounding nuclei (STN, SNr, RN) are not directly hit by EC, indicating a predominant white matter stimulation. The individual EC position within the triangle cannot be predicted and is based on individual slMFB (tractography) geometry. There was one intracranial bleeding (FORESEE I study) during a first implantation attempt in a patient who later received full bilateral implantation. Typical oculomotor side effects are idiosyncratic for the target region and at inferior contacts. Conclusion: The detailed surgical procedure of slMFB DBS implantation has not been described before. The slMFB emerges as an interesting region for the treatment of major depression (and other psychiatric diseases) with DBS. So far it has only been successfully researched in open label clinical case series and in 15 patients published. Stimulation probably achieves its effect through direct white-matter modulation of slMFB fibers. The surgical implantation comprises a standardized protocol combining tractographic imaging based on DTI, targeting and electrophysiological evaluation of the target region. To this end, slMFB DBS surgery is in technical aspects comparable to typical movement disorder surgery. In our view, slMFB DBS should only be performed under tractographic assistance.

Tractography Study of Deep Brain Stimulation of the Anterior Cingulate Cortex in Chronic Pain: Key to Improve the Targeting.

BACKGROUND: Deep brain stimulation (DBS) of the anterior cingulate cortex (ACC) is a new treatment for alleviating intractable neuropathic pain. However, it fails to help some patients. The large size of the ACC and the intersubject variability make it difficult to determine the optimal site to position DBS electrodes. The aim of this work was therefore to compare the ACC connectivity of patients with successful versus unsuccessful DBS outcomes to help guide future electrode placement. METHODS: Diffusion magnetic resonance imaging (dMRI) and probabilistic tractography were performed preoperatively in 8 chronic pain patients (age 53.4 ± 6.1 years, 2 females) with ACC DBS, of whom 6 had successful (SO) and 2 unsuccessful outcomes (UOs) during a period of trialing. RESULTS: The number of patients was too small to demonstrate any statistically significant differences. Nevertheless, we observed differences between patients with successful and unsuccessful outcomes in the fiber tract projections emanating from the volume of activated tissue around the electrodes. A strong connectivity to the precuneus area seems to predict unsuccessful outcomes in our patients (UO: 160n/SO: 27n), with (n), the number of streamlines per nonzero voxel. On the other hand, connectivity to the thalamus and brainstem through the medial forebrain bundle (MFB) was only observed in SO patients. CONCLUSIONS: These findings could help improve presurgical planning by optimizing electrode placement, to selectively target the tracts that help to relieve patients' pain and to avoid those leading to unwanted effects.

Ventral tegmental area dopaminergic lesion-induced depressive phenotype in the rat is reversed by deep brain stimulation of the medial forebrain bundle.

DBS of the medial forebrain bundle (MFB) has been investigated clinically in major depressive disorder patients with rapid and long-term reduction of symptoms. In the context of chronic bilateral high frequency deep brain stimulation (DBS) of the MFB, the current study looked at the impact of lesioning the ascending dopaminergic pathway at the level of the ventral tegmental area (VTA). Sprague-Dawley female rats were given bilateral injection of 6-OHDA into the VTA (VTA-lx group) or were left unlesioned (control group). Later, all animals received bilateral microelectrode implantation into the MFB followed by chronic continuous stimulation for 3 weeks. Behavioral tests were performed as baseline and following MFB-DBS, along with histological analysis. Pre-stimulation baseline testing of the VTA-lx animals indicated depressive-like phenotype in comparison with controls. Response to MFB-DBS varied according to (i) the degree of dopaminergic depletion: animals with severe mesocorticolimbic dopamine depletion did not, whilst those with mild dopamine loss responded well to stimulation; (ii) environmental conditions and the nature of the behavioral tests, e.g., stressful vs non-stressful situations. Neuromodulation-induced c-fos expression in the prelimbic frontal cortex and nucleus accumbens was also dependent upon integrity of the dopaminergic ascending projections. Our results confirm a potential role for dopamine in symptom relief observed in clinical MFB-DBS. Although mechanisms are not fully understood, the data suggests that the rescue of depressive phenotype in rodents can work via both dopamine-dependent and independent mechanisms. Further investigations concerning the network of depression using neuromodulation platforms in animal models might give insight into genesis and treatment of major depression disorder.

Feasibility and safety of continuous and chronic bilateral deep brain stimulation of the medial forebrain bundle in the naïve Sprague-Dawley rat.

OBJECTIVE: Deep brain stimulation (DBS) of the superolateral branch of the medial forebrain bundle (MFB) has provided rapid and dramatic reduction of depressive symptoms in a clinical trial. Early intracranial self-stimulation experiments of the MFB suggested detrimental side effects on the animals' health; therefore, the current study looked at the viability of chronic and continuous MFB-DBS in rodents, with particular attention given to welfare issues and identification of stimulated pathways. METHODS: Sprague-Dawley female rats were submitted to stereotactic microelectrode implantation into the MFB. Chronic continuous DBS was applied for 3-6 weeks. Welfare monitoring and behavior changes were assessed. Postmortem histological analysis of c-fos protein expression was carried out. RESULTS: MFB-DBS resulted in mild and temporary weight loss in the animals, which was regained even with continuing stimulation. MFB-DBS led to increased and long-lasting c-fos expression in target regions of the mesolimbic/mesocortical system. CONCLUSIONS: Bilateral continuous chronic MFB-DBS is feasible, safe, and without impact on the rodent's health. MFB-DBS results in temporary increase in exploration, which could explain the initial weight loss, and does not produce any apparent behavioral abnormalities. This platform represents a powerful tool for further preclinical investigation of the MFB stimulation in the treatment of depression.

Transcriptional and structural plasticity of tyrosine hydroxylase expressing neurons in both striatum and nucleus accumbens following dopaminergic denervation.

Mice that express green fluorescent protein (GFP) under the control of tyrosine hydroxylase (TH) gene promoter were used to visualize transcriptional as well as structural regulation of TH cells following prolonged dopaminergic denervation. A unilateral lesion of the medial forebrain bundle was induced by 6-hydroxydopamine. In the unlesioned contralateral striatum and nucleus accumbens surprisingly high numbers of resident GFP-positive neurons (about 2653 and 422 per striatum and accumbens, respectively) were observed while only much lower TH-positive neurons (about 214 and 102 per striatum and accumbens, respectively) were detectable. In the lesioned hemisphere the number of GFP neurons was slightly increased already at day 4 by 16% and more at day 40 by 47% while the number of TH-immunoreactive neurons was dramatically increased by 848% at day 4 and by 1139% at day 40 over the control side. Additionally and particularly pronounced in the nucleus accumbens, GFP-positive neurons demonstrated increased sprouting of their projections over time, stronger than observed by TH-immunostaining. The load in TH protein may be essentially determined by post-transcriptional suppression/degradation while GFP may rather reflect the gross transcriptional activity. Thus, permanent dopaminergic pathway injury induces both transcriptional as well as structural plasticity of TH expressing neurons in striatal and accumbal target areas of ventral midbrain dopaminergic neurons.

Fascículos asociativos ínsulo-operculares: revisión de su anatomía y de las implicaciones para el abordaje transopercular a la ínsula / Insular-opercular associative tracts: Review of their anatomy and relevance for the trans-opercular approach to the insula

INTRODUCCIÓN: No hay estudios que revisen en detalle y de forma sistemática la anatomía de los fascículos asociativos ínsulo-operculares. OBJETIVO: En el presente trabajo, se realizó una extensa revisión de la literatura reciente de las fibras de asociación relacionadas con el lóbulo de la ínsula y los opérculos. RESULTADOS: Los tractos conectados con los opérculos son el fascículo frontal oblicuo, el fascículo arcuato, la porción horizontal del fascículo longitudinal superior y el fascículo longitudinal medial. A nivel de la ínsula, el fascículo fronto-occipital inferior (FFOI) discurre paralelo al fascículo uncinado, atravesando la porción antero-inferior de la cápsula externa y el claustrum. CONCLUSIONES: La pars triangular y orbicular de la circunvolución frontal inferior y la parte media y anterior de la circunvolución temporal superior están menos conectadas con la red perisilviana asociativa. De esta forma, constituyen 2 corredores anatómicos para el abordaje transopercular a la ínsula

INTRODUCTION: The insula is a highly connected area, as an intricate network of afferent and efferent projections connect it with adjacent and distant cortical regions. Objective: To perform an extensive review of recent literature to analyse the anatomy of the associative tracts related to the insula. RESULTS: The frontal aslant tract, arcuate fasciculus, horizontal portion of the superior longitudinal fasciculus and the middle longitudinal fasciculus are associative tracts connected to the opercula. The inferior fronto-occipital fasciculus (IFOF) and uncinate fasciculus run under the anterior and inferior portion of the insula. CONCLUSIONS: the pars triangularis and orbicularis of the inferior frontal gyrus, as well as the middle and anterior part of the superior temporal gyrus, have few connections with the perisylvian associative network. Consequently, in the trans-opercular approach to the insula, these 2 regions represent anatomical corridors that give access to the insula. The IFOF and the uncinate fasciculus represent the deep functional margin of resection

Intrastriatal excitotoxic lesion or dopamine depletion of the neostriatum differentially impairs response execution in extrapersonal space.

Dysfunction of the neostriatum, a primary feature of several neurodegenerative disorders, including Parkinson's disease and Huntington's disease, has been found to result in impaired localisation of, and reaction to, contralateral stimuli. On the basis of previous findings, it is hypothesised that, with increasing eccentricity of the response option, striatal cell loss may impair response localisation at the furthest levels of eccentricity, whereas dopamine (DA) depletion may not impact adversely upon responses executed in this extrapersonal space. In order to elucidate more fully the function of the striatum, the present study examined the differential impact of unilateral DA depletion or excitoxic lesion on response execution in ipsilateral and contralateral space at up to four levels of eccentricity. The results confirmed that, after both types of striatal dysfunction, the sensory ability to detect stimuli remains intact, whereas the ability to direct responses in absolute contralateral space is impaired. Distinct differences in the profiles of impairment were, however, evident, with a marked increase in response omissions observed after DA depletion, which may reflect decreased motivational processing, and recovery of function observed in rats with excitotoxic lesions, which suggests the ability to re-learn. Furthermore, the data demonstrate that, after cell loss, responding in near contralateral space is controlled by competing striata, whereas responding in extrapersonal space relies on the contralateral hemisphere. These results have implications for understanding the role of the striatum in egocentrically defined response localisation, as well as for unravelling the behavioural impact of striatal cell loss or aberrant DA transmission observed in neurodegenerative diseases.

Increased efficacy of the 6-hydroxydopamine lesion of the median forebrain bundle in small rats, by modification of the stereotaxic coordinates.

The 6-hydroxydopamine (6-OHDA) lesion is the most widely used rat model of Parkinson's disease. A single unilateral injection of 6-OHDA into the median forebrain bundle (MFB) selectively destroys dopamine neurons in the ipsilateral substantia nigra pars compacta (SNc) and ventral tegmental area (VTA), removing more than 95% of the dopamine innervation from target areas. The stereotaxic coordinates used to deliver 6-OHDA to the MFB have been used in our laboratory successfully for more than 25 years. However, in recent years we have observed a decline in the success rate of this lesion. Previously regular success rates of >80% of rats lesioned, have become progressively more variable, with rates as low as 20% recorded in some experiments. Having excluded variability of the neurotoxin and operator errors, we hypothesized that the change seen might be due to the use of smaller rats at the time of first surgery. An attempt to proportionally adjust the lesion coordinates base on head size did not increase lesion efficacy. However, in support of the small rat hypothesis it was observed that, using the standard coordinates, rat's heads had a "nose-up" position in the stereotaxic fame. Adjustment of the nose bar to obtain a flat head position during surgery improved lesion success, and subsequent adjustments of the lesion coordinates to account for smaller head size led to a greatly increased lesion efficacy (>90%) as assessed by amphetamine induced rotation.

Riluzole improves motor deficits and attenuates loss of striatal neurons in a sequential double lesion rat model of striatonigral degeneration (parkinson variant of multiple system atrophy).

We investigated neuroprotective effects of riluzole, an anti-glutamatergic agent that is FDA approved for disease-modifying therapy in amyotrophic lateral sclerosis (ALS), in an established double lesion rat model of striatonigral degeneration (SND), the neuropathological substrate of parkinsonism associated with MSA (MSA-P). Riluzole was administered prior to and consecutively for ten days following double lesion placement in the left-sided medial forebrain bundle and ipsilateral striatum. Assessment of motor behaviour using a flex field system showed a significant reduction of motor disturbance in animals with striatonigral lesions treated with riluzole compared to lesioned but untreated animals (P<0.001). DARPP-32 immunohistochemistry revealed a significant reduction of absolute striatal lesion volume in riluzole treated animals compared to lesioned but untreated animals (P<0.01). No significant difference in counts of nigral dopaminergic neurons was found in treated versus untreated double-lesioned animals. The results of our study indicate that riluzole mediates neuroprotective effects in the double lesion rat model of MSA-P. Whether riluzole also protects autonomic and cerebellar pathways that are frequently affected in MSA remains to be determined. Nonetheless, our study is the first to provide an experimental rationale for exploring possible neuroprotective effects of riluzole in MSA.

Electro-acupuncture stimulation protects dopaminergic neurons from inflammation-mediated damage in medial forebrain bundle-transected rats.

Through producing a variety of cytotoxic factors upon activation, microglia are believed to participate in the mediation of neurodegeneration. Intervention against microglial activation may therefore exert a neuroprotective effect. Our previous study has shown that the electro-acupuncture (EA) stimulation at 100 Hz can protect axotomized dopaminergic neurons from degeneration. To explore the underlying mechanism, the effects of 100 Hz EA stimulation on medial forebrain bundle (MFB) axotomy-induced microglial activation were investigated. Complement receptor 3 (CR3) immunohistochemical staining revealed that 24 sessions of 100 Hz EA stimulation (28 days after MFB transection) significantly inhibited the activation of microglia in the substantia nigra pars compacta (SNpc) induced by MFB transection. Moreover, 100 Hz EA stimulation obviously inhibited the upregulation of the levels of tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta mRNA in the ventral midbrains in MFB-transected rats, as revealed by reverse transcriptase polymerase chain reaction (RT-PCR). ED1 immunohistochemical staining showed that a large number of macrophages appeared in the substantia nigra (SN) 14 days after MFB transection. The number of macrophages decreased by 47% in the rats that received 12 sessions of EA simulation after MFB transection. These data indicate that the neuroprotective role of 100 Hz EA stimulation on dopaminergic neurons in MFB-transected rats is likely to be mediated by suppressing axotomy-induced inflammatory responses. Taken together with our previous results, this study suggests that the neuroprotective effect of EA on the dopaminergic neurons may stem from the collaboration of its anti-inflammatory and neurotrophic actions.

Microglial phagocytosis of dopamine neurons at early phases of apoptosis.

Transection of the medial forebrain bundle caused apoptosis of dopamine neurons in the rat substantia nigra. Immunohistochemical localization of activated microglia and tyrosine hydroxylase in the axotomized substantia nigra showed that activation of microglia was rapid and OX-6 (MHC-II marker)-positive and ED1 (lysosomal phagocytic marker)-positive microglia were apposed to structurally intact tyrosine hydroxylase-positive dopamine neurons, indicating microglial phagocytosis of degenerating dopamine neurons. The occurrence of microglial phagocytosis at early stages of apoptosis may indicate the evolution of apoptosis into an irreversible state. Alternatively, interventions that suppress early activation of microglia might lead to novel mechanisms for neuron protection.

Long-term high-frequency electro-acupuncture stimulation prevents neuronal degeneration and up-regulates BDNF mRNA in the substantia nigra and ventral tegmental area following medial forebrain bundle axotomy.

Electroacupuncture (EA) has been used in China for many years to treat Parkinson's disease (PD) with reportedly effective results. However, the physiological and biological mechanism behind its effectiveness is still unknown. In the present study, different frequencies of chronic EA stimulation (0, 2, 100 Hz) were tested in a partially lesioned rat model of PD which was induced by transection of the medial forebrain bundle (MFB). After 24 sessions of EA stimulation (28 days after MFB transection), dopaminergic neurons in the ventral midbrain were examined by immunohistochemical staining, and brain-derived neurotrophic factor (BDNF) mRNA levels in ventral midbrain were measured by in situ hybridization. The results show a marked decrease of dopaminergic neurons on the lesioned side of the substantia nigra (SN) comparing with the unlesioned side. Zero Hz and 2 Hz EA stimulation had no effect on the disappearance of dopaminergic neurons. However, after 100 Hz EA, about 60% of the tyrosine hydroxylase (TH)-positive neurons remained on the lesioned side of the SN. In addition, levels of BDNF mRNA in the SN and ventral tegmental area (VTA) of the lesioned side were significantly increased in the 100 Hz EA group, but unchanged in the 0 and 2 Hz groups. Our results suggest that long-term high-frequency EA is effective in halting the degeneration of dopaminergic neurons in the SN and up-regulating the levels of BDNF mRNA in the subfields of the ventral midbrain. Activation of endogenous neurotrophins by EA may be involved in the regeneration of the injured dopaminergic neurons, which may underlie the effectiveness of EA in the treatment of PD.

Amyloid precursor protein gene disruption attenuates degeneration of substantia nigra compacta neurons following axotomy.

Our past work has shown that the C-terminal fragment of amyloid precursor protein (APP) translocated to the nucleus in neurons destined for delayed excitotoxic degeneration. To test whether nuclear APP fragments also play a role in the progressive loss of dopaminergic (DA) substantia nigra compacta (SNc) neurons, we performed unilateral medial forebrain bundle (MFB) transection on APP wild type (WT) and on mice with disruption of the APP gene (KO). In WT mice immunoreactivity for APP C-terminal, beta-amyloid and Alz90 epitopes appeared in the nuclei of axotomized DA neurons at 3 days post-lesion (dpl), persisted at 7 dpl and was absent in 14 dpl mice. APP N-terminal immunoreactivity was restricted to the cytosol at all time points, precluding the possibility of full length APP in the nucleus. Nuclear localization of APP epitopes was absent in neurons of the contralateral SNc or in neurons of the ipsilateral ventral tegmental area and SN reticulata. The presence of APP C-terminal and Alz90 domains was confirmed by Western blotting performed on the nuclear fraction of the SN ipsilateral to the axotomy. Quantitative morphometric analysis revealed that WT mice demonstrated earlier and more profound loss of tyrosine hydroxylase+SNc neurons than did KO mice. These data showed that a novel nuclear C-terminal fragment appeared coincident with SNc neuron degeneration, and that APP deficiency correlated with significant neuroprotection in vivo.

Responses of ventral tegmental area GABA neurons to brain stimulation reward.

Dopamine neurons in the ventral tegmental area (VTA) have been implicated in rewarded behaviors, including intracranial self-stimulation (ICSS). We demonstrate, in unrestrained rats, that the discharge activity of a homogeneous population of presumed VTA GABA neurons, implicated in cortical arousal, increases before ICSS of the medial forebrain bundle (MFB). These findings suggest that VTA GABA neurons may be involved in the attentive processes related to brain stimulation reward (BSR).

Upregulation of BDNF mRNA and trkB mRNA in the nigrostriatal system and in the lesion site following unilateral transection of the medial forebrain bundle.

We have performed unilateral transection of the medial forebrain bundle (MFB) and studied BDNF mRNA and trkB mRNA levels at different postlesion times in the nigrostriatal system by means of in situ hybridization. BDNF mRNA levels were transiently induced in the substantia nigra pars compacta at 1 day postaxotomy. The disposition of BDNF mRNA expressing cells at this postlesion time in substantia nigra mimicked that of the dopaminergic neurons expressing the mRNA for the dopamine transporter. TrkB mRNA levels remained unaltered in the ventral mesencephalon at the different postlesion times examined-1 to 14 days. In contrast, trkB mRNA levels were significantly induced in the striatum at the longer postlesion time examined-14 days-when all neurodegenerative events are completed. It is becoming apparent that nigral BDNF mRNA levels are anterogradely transported to its target tissue in striatum. However, following axotomy, the lesion site represents a second potential target for BDNF action. Consequently, we also analyzed the pattern of mRNA expression for BDNF and trkB at the lesion site where dopaminergic axons are disconnected. There, we found notable inductions of both BDNF mRNA and trkB mRNA levels at 4 days postaxotomy. BDNF mRNA expressing cells were confined at the site of axotomy, which coincided precisely to that showing induction of trkB mRNA. Altogether, our results anticipate promising trophic roles of BNDF in the injured nigrostriatal system.

Robust regeneration of CNS axons through a track depleted of CNS glia.

Transected CNS axons do not regenerate spontaneously but may do so if given an appropriate environment through which to grow. Since molecules associated with CNS macroglia are thought to be inhibitory to axon regeneration, we have tested the hypothesis that removing these cell types from an area of brain will leave an environment more permissive for axon regeneration. Adult rats received unilateral knife cuts of the nigrostriatal tract and ethidium bromide (EB) was used to create a lesion devoid of astrocytes, oligodendrocytes, intact myelin sheaths, and NG2 immunoreactive cells from the site of the knife cut to the ipsilateral striatum (a distance of 6 mm). The regenerative response and the EB lesion environment was examined with immunostaining and electron microscopy at different timepoints following surgery. We report that large numbers of dopaminergic nigral axons regenerated for over 4 mm through EB lesions. At 4 days postlesion dopaminergic sprouting was maximal and the axon growth front had reached the striatum, but there was no additional growth into the striatum after 7 days. Regenerating axons did not leave the EB lesion to form terminals in the striatum, there was no recovery of function, and the end of axon growth correlated with increasing glial immunoreactivity around the EB lesion. We conclude that the removal of CNS glia promotes robust axon regeneration but that this becomes limited by the reappearance of nonpermissive CNS glia. These results suggest, first, that control of the glial reaction is likely to be an important feature in brain repair and, second, that reports of axon regeneration must be interpreted with caution since extensive regeneration can occur simply as a result of a major glia-depleting lesion, rather than as the result of some other specific intervention.

Brain-stimulation reward thresholds raised by an antisense oligonucleotide for the M5 muscarinic receptor infused near dopamine cells.

Oligonucleotides targeting M5 muscarinic receptor mRNA were infused for 6 d into the ventral tegmental area of freely behaving rats trained to bar-press for lateral hypothalamic stimulation. The bar-pressing rate was determined at a range of frequencies each day to evaluate the effects of infusions on reward. M5 antisense oligonucleotide (oligo) infusions increased the frequency required for bar pressing by 48% over baseline levels, with the largest increases occurring after 4-6 d of infusion. Two control oligos had only slight effects (means of 5 and 11% for missense and sense oligos, respectively). After the infusion, the required frequency shifted back to baseline levels gradually over 1-5 d. Antisense oligo infusions decreased M5 receptors on the ipsilateral, but not the contralateral, side of the ventral tegmentum, as compared with a missense oligo. Therefore, M5 muscarinic receptors associated with mesolimbic dopamine neurons seem to be important in brain-stimulation reward.

Angiotensin II receptor binding sites in the ventral portion of the bed nucleus of the stria terminalis are reduced by interruption of the medial forebrain bundle.

Many techniques have been utilized to discern the localization of angiotensin II (Ang II) receptors to specific cellular components (glia, neuronal cell bodies and nerve terminals) in the brain. In the present study, we used lesioning techniques to localize Ang II receptors to cellular components in the rat forebrain. In the first experiment, axons ascending to the hypothalamus and forebrain from neurons in the brainstem were destroyed by unilaterally cutting the medial forebrain bundle (MFB). In the second experiment, a single injection of the neurotoxin, ibotenic acid, was injected unilaterally into the ventral portion of the bed nucleus of the stria terminalis (BSTV) to destroy neuronal cell bodies, thus determining if Ang II receptors are present on neuronal cell bodies. In both experiments, the animals were sacrificed after two weeks recovery and the brains processed for in vitro receptor autoradiography using 125I-sar1,ile8 Ang II (125I-SI Ang II). Unilateral knife-cut lesions of the MFB caused a significant reduction in 125I-SI Ang II binding in the BSTV (30+/-6%) and the piriform cortex (PC; 26+/-4%) ipsilateral to the knife cut. Unilateral injection of the neurotoxin into the BSTV failed to alter 125I-SI Ang II binding in this nucleus. These experiments suggest that at least a subpopulation of Ang II receptors in the BSTV and PC are located on terminals of neurons that have their cell bodies in the brainstem and their axons in the MFB.