Percutaneous nerve field stimulation (PENS) of the occipital region as a possible predictor for occipital nerve stimulation (ONS) responsiveness in refractory headache disorders? A feasibility study.

BACKGROUND: Occipital nerve stimulation (ONS) has been reported to diminish pain levels in intractable chronic headache syndromes of different origin. No reliable objective markers exist to predict ONS responsiveness. This study investigated the predictive value of occipital percutaneous nerve field stimulation (PENS) prior to ONS. METHODS: This trial included 12 patients (CCH, CM, PTH, CH) with chronic refractory headache syndromes eligible for ONS. Repetitive PENS (3 × /10 days) was performed and the headache severity/frequency monitored over four weeks before ONS implantation. Further assessment of PENS/ONS outcomes were stimulation-related complications, perception/tolerance stimulation threshold, the Migraine Disability Scale (MIDAS) and the Beck Depression Inventory (BDI). RESULTS: All PENS responders benefited from ONS. Of the seven PENS-nonresponders with VAS 6.1(±1.1), six experienced significant pain relief from ONS after three months and one patient failed the PENS/ONS trial (VAS 3.7 (±1.6)); (95% CI 3.6 to 5.7, p < 0.001). The VAS baseline was 8.4 (±0.5) and decreased significantly (50% reduction in severity/frequency) in five patients after PENS, while seven failed to improve (VAS 4.9 (±1.1); (95% CI 2.5 to 4.5, p < 0.001). BDI baseline (from 22.6 (±4.2) to 10.6 (±5.9) (95% CI 7.4 to 16.6, p < 0.001)) and MIDAS baseline (from 143.9 (±14.5) to 72.8 (±28.7) (95% CI 1.17 to 2.3, p < 0.001)) significantly declined after ONS. No PENS/ONS-related complications occurred. CONCLUSIONS: Presurgical applied occipital PENS failed to identify ONS responders sufficiently according to our study protocol, thus requiring further specific investigations to determine its predictive usefulness.

The impact of brain shift in deep brain stimulation surgery: observation and obviation.

BACKGROUND: The impact of brain shift on deep brain stimulation surgery is considerable. In DBS surgery, brain shift is mainly caused by CSF loss. CSF loss can be estimated by post-surgical intracranial air. Different approaches and techniques exist to minimize CSF loss and hence brain shift. The aim of this survey was to investigate the extent and dynamics of CSF loss during DBS surgery, analyze its impact on final electrode position, and describe a simple and inexpensive method of burr hole closure. METHODS: Sixty-six patients being treated with deep brain stimulation were retrospectively analyzed for this treatise. During surgery, CSF loss was minimized using bone wax as a burr hole closure. Intracranial air volume was calculated based on early post-surgery stereotactic 3D CT and correlated with duration of surgery and electrode deviations derived from post-surgery image fusion. RESULTS: Median early post-surgery intracranial air was 2.1 cm(3) (range 0-35.7 cm(3), SD 8.53 cm(3)). No correlation was found between duration of surgery and CSF-loss (R = 0.078, p = 0.534), indicating that CSF loss mainly occurs early during surgery. Linear regression analysis revealed no significant correlations regarding volume of intracranial air and electrode displacement in any of the three principal axes. No significant difference regarding electrode deviations between first and second side of surgery were observed. CONCLUSIONS: CSF loss mainly occurs during the early phase of DBS surgery. CSF loss during a later phase of surgery can be effectively averted by burr hole closure. Postoperative intracranial air volumes up to 35 cm(3) did not result in significant electrode displacement in our series. Comparing our results to studies previously published on this subject, burr hole closure using bone wax is highly effective.

GPi-DBS may induce a hypokinetic gait disorder with freezing of gait in patients with dystonia.

OBJECTIVES: Stimulation-induced hypokinetic gait disorders with freezing of gait (FOG) have been reported only recently as adverse effects of deep brain stimulation (DBS) of the globus pallidus internus (GPi) in patients with dystonia. The aim of this work was to determine the frequency and the nature of this GPi-DBS-induced phenomenon. METHODS: We retrospectively screened our database of patients with dystonia who underwent DBS. Patients with focal, segmental, or generalized dystonia of primary or tardive origin and no gait disorder due to lower limb dystonia before DBS, bilateral pallidal stimulation, and a follow-up for more than 6 months were included. Reports of adverse events were analyzed, and gait abnormalities were scored by comparing preoperative and postoperative video recordings using Movement Disorder Society-sponsored revision of the Unified Parkinson's Disease Rating Scale (MDS-UPDRS) items 3.10 (gait) and 3.11 (FOG). To assess the role of GPi-DBS in gait abnormalities, DBS was paused for 24 hours. Gait and FOG were assessed 30 minutes, 2 hours, and 24 hours after restarting DBS. Finally, a standardized adjustment algorithm was performed trying to eliminate the gait disorder. RESULTS: Of a collective of 71 patients with dystonia, 6 presented with a new gait disorder (8.5%; 2 men, 4 women, mean age 61.3 years [48-69 years], 2 craniocervical, 1 DYT-1 segmental, 1 truncal, 2 tardive dystonia). GPi-DBS improved Burke-Fahn-Marsden Dystonia Rating Scale motor score by 54% and disability score by 52%. MDS-UPDRS item 3.10 worsened from 0.5 (±0.8) to 2.0 (±0.9) and item 3.11 from 0 to 2.5 (±0.5). The gait disorder displayed shuffling steps and difficulties with gait initiation and turning. Increasing voltages improved dystonia but triggered FOG, sometimes worsening over a period of a few hours. It vanished within minutes after ceasing DBS. Electrode misplacement was ruled out. In all but one patient, no optimal configuration was found despite extensive testing of settings (monopolar, bipolar, pulse width 60-210 µs, frequency 60-180 Hz). Nevertheless, a compromise between optimal stimulation for dystonia and eliciting FOG was achieved in each case. CONCLUSIONS: A hypokinetic gait disorder with FOG can be a complication of GPi-DBS.

Battery lifetime in pallidal deep brain stimulation for dystonia.

BACKGROUND AND PURPOSE: The aim of the study was to analyse the lifetime of Soletra implantable pulse generators (IPG) in deep brain stimulation (DBS) of the globus pallidus internus (GPi) for dystonia, depending on stimulation parameters and the total electrical energy delivered (TEED) by the IPG. METHODS: In a prospective series of 20 patients with GPi DBS for dystonia, we recorded IPG longevity and stimulation parameters over time. An evaluation of the TEED was performed using the previously suggested equation [(voltage(2) × pulse width × frequency)/impedance] × 1 s. RESULTS: During median follow-up of 57 months (range 23-79 months), 64 IPGs were replaced because of battery depletion or end of life signal. We found a mean IPG longevity of 25.1 ± 10.1 (range 16-60) months, which was inversely correlated with the TEED (r = -0.72; P < 0.001). IPG longevity was not different between bipolar and monopolar stimulation (24.9 ± 10.8 vs. 25.4 ± 9.0 months, P = 0.76). Incongruously, the mean TEED applied throughout the lifetime cycle was significantly higher in patients with bipolar compared with monopolar stimulation (584 ± 213 vs. 387 ± 121 Joule; P < 0.01). CONCLUSIONS: Battery lifetime in GPi DBS for dystonia is substantially shorter compared with that reported in DBS for Parkinson's disease, caused by a considerably higher voltage and greater pulse width and therefore a higher TEED applied during the battery lifetime cycle. The commonly used equation to calculate TEED, however, seems to be correct only for monopolar, but not bipolar stimulation.

Neonatal lesions of the entorhinal cortex induce long-term changes of limbic brain regions and maze learning deficits in adult rats.

We here investigated the effects of neonatal lesions of the entorhinal cortex (EC) in rats on maze learning and on structural alterations of its main projection region, the hippocampus, as well as other regions with anatomical connections to the EC that are involved in maze learning. Since early brain damage is considered to be involved in certain neuropsychiatric diseases, this approach sought to model certain aspects of this etiopathogenesis. Bilateral neonatal lesions were induced on postnatal day 7 by microinjection of ibotenic acid (1.3 microg/0.2 microl phosphate-buffered saline (PBS)) into the EC. Naive and sham-lesioned rats served as controls. Rats were trained and tested on an eight-arm radial maze for allocentric and egocentric learning. Subsequently, gold-chloride staining and immunohistochemical staining for the microtubule-associated protein MAP-2 was used to assess myelination and dendritic density in the hippocampus, striatum and medial prefrontal cortex (mPFC) of these rats. Additionally, parvalbumin-expressing, presumably GABAergic interneurons, were evaluated in these regions. Performance in both the allocentric and the egocentric strategy was disturbed after neonatal EC lesion as shown by an increase of repeated arm entries, which indicates disturbed working memory. Histological evaluation revealed that the density of parvalbumin-immunopositive neurons and myelin sheaths was reduced in the hippocampus but not in the striatum and mPFC in neonatally lesioned rats. Density of MAP-2 staining did not differ between groups in all regions tested. Since structural alterations were only found in the EC and hippocampus our findings support their eminent role in working memory and show that no functional restoration occurs after neonatal lesions.

Hypothalamic deep brain stimulation for cluster headache: experience from a new multicase series.

Deep brain stimulation (DBS) of the posterior hypothalamus was found to be effective in the treatment of drug-resistant chronic cluster headache. We report the results of a multicentre case series of six patients with chronic cluster headache in whom a DBS in the posterior hypothalamus was performed. Electrodes were implanted stereotactically in the ipsilateral posterior hypothalamus according to published coordinates 2 mm lateral, 3 mm posterior and 5 mm inferior referenced to the mid-AC-PC line. Microelectrode recordings at the target revealed single unit activity with a mean discharge rate of 17 Hz (range 13-35 Hz, n = 4). Out of six patients, four showed a profound decrease of their attack frequency and pain intensity on the visual analogue scale during the first 6 months. Of these, one patient was attack free for 6 months under neurostimulation before returning to the baseline which led to abortion of the DBS. Two patients had experienced only a marginal, non-significant decrease within the first weeks under neurostimulation before returning to their former attack frequency. After a mean follow-up of 17 months, three patients are almost completely attack free, whereas three patients can be considered as treatment failures. The stimulation was well tolerated and stimulation-related side-effects were not observed on long term. DBS of the posterior inferior hypothalamus is an effective therapeutic option in a subset of patients. Future controlled multicentre trials will need to confirm this open-label experience and should help to better define predictive factors for non-responders.

Immunostaining for cell picking and real-time mRNA quantitation.

Microdissection techniques allow a cell-type or even cell-specific mRNA analysis within complex tissues. Furthermore, valid mRNA quantitation can be performed by real-time reverse transcriptase-polymerase chain reaction from a few isolated cells obtained from cryosections. For a more precise access to many cell types, this technique has to be complemented by a cell-type-specific immunostaining. To evaluate its effect on mRNA quantitation, we analyzed alveolar macrophages (AMs) from control rat lungs and those undergoing stimulation with lipopolysaccharide and interferon-gamma nebulization. Whereas AMs from the left lung were directly harvested for mRNA extraction by bronchoalveolar lavage, tissue sections of the right lung were stained with an optimized immunofluorescence protocol detecting AMs. Fifteen AM profiles per sample were picked by laser-assisted sampling technique. Normalizing to a standard gene, nitric oxide synthase II (NOSII) and tumor necrosis factor (TNF)-alpha mRNA were quantified by real-time reverse transcriptase-polymerase chain reaction. In stimulated lungs, the percentage of picked samples positive for NOSII or TNF-alpha mRNA increased significantly. Moreover, a marked increase in the ratio of target gene mRNA to standard gene mRNA was noted for both NOSII and TNF-alpha in picked AMs from stimulated lungs, which matched very well the increase detected in the lavaged AMs undergoing direct RNA extraction. Thus, when using an optimized protocol for immunofluorescence, this approach may be reliably combined with laser-assisted cell picking and real-time mRNA quantitation in a few immunohistochemically characterized cell profiles within complex tissues.

Immunostaining and laser-assisted cell picking for mRNA analysis.

Isolation of single cells or cell clusters from complex tissue sections has become possible by microdissection techniques. Employing laser-assisted cell picking, cell-specific mRNA analysis of a few isolated cell profiles may be performed. However, microscopic discrimination of different cell types in routinely stained tissue sections is limited, whereas immunostaining enables a more precise access to cells of interest. This approach was noted to interfere with mRNA recovery. To define optimal conditions for mRNA amplification from immunodetected cells, we systematically investigated several potential affectors. Kind of fixation, antibodies and staining reagents, incubation and total processing time, and digestion with proteinase K turned out to influence mRNA stability. We present rapid protocols for immunohistochemistry and immunofluorescence with total incubation times of approximately 25 to 40 minutes and 10 to 20 minutes, respectively, and suggest mRNA amplification without a preceding extraction step. Applying these protocols to oligocellular clusters containing approximately 20 cell profiles and nuclei each from lung and kidney tissue, the highest efficiency rates of mRNA amplification were obtained when combining short-term formalin fixation, reduction of antibody incubation time, application of immunofluorescence, and digestion with proteinase K. Thus, the successful combination of immunostaining and laser-assisted cell picking remarkably improves cell type-specific analysis of gene expression within complex tissues.

Cell type-specific mRNA quantitation in non-neoplastic tissues after laser-assisted cell picking.

OBJECTIVE: Cell type-specific mRNA quantitation can be reliably performed after harvesting less than 20 cell profiles from haemalaun-stained cryosections by laser-assisted cell picking. Up to now it has been unclear to what extent these techniques can be used to analyze differential gene expression in complex tissues. METHODS: Using a rat model of experimental endotoxin priming of the lung various pulmonary cell types were microdissected from isolated perfused and ventilated rat lungs after aerosol lipopolysaccharide/interferon-gamma stimulation. RESULTS: Porphobilinogen deaminase housekeeping gene (PBGD) and nitric oxide synthase II (NOSII) mRNA in arterial endothelial cells (AEC), bronchiolar epithelial cells (BEC), alveolar septum containing monocytes/macrophages (AS+), alveolar septum without monocytes/macrophages (AS-) and intraluminar alveolar macrophages (AM) could be quantified by real-time RT-PCR. The strongest upregulation of NOSII mRNA occurred in AM, while minimal NOSII expression was detected in BEC, AS+ and AS-. In AEC NOSII mRNA was not detectable. CONCLUSION: The combination of laser microdissection and real-time RT-PCR is a valuable tool for the quantitative in situ characterization of differential gene expression within complex tissues.