Technique of injection of onabotulinumtoxin A for chronic migraine: the PREEMPT injection paradigm.

In 2013 the Italian Pharmacy Agency (AIFA) approved onabotulinumtoxin A injection to prevent headaches in adult patients with chronic migraine (headaches on at least 15 days per month of which at least 8 days are with migraine) that has not responded to at least three prior pharmacological prophylaxis therapies and whose condition is appropriately managed for medication overuse. In the present paper we report the method of injection of Onabotulinumtoxin A for chronic migraine based on the PREEMPT paradigm as described by Blumenfeld et al. (Headache 50:1406-1418, 2010) adapted to our clinical setting.

Hypertension is a factor associated with chronic daily headache.

Chronic daily headache (CDH) is one of the more frequently observed headache syndromes at major tertiary care centers. CDH is defined as headache occurring >15 days/month. Different mechanisms are involved in the development of CDH but what factors specifically contributing to the transformation from episodic into CDH remain largely unknown. Analgesic overuse is commonly identified as the most important factor for such transformation. Hypertension, allergy, asthma, arthritis, diabetes, obesity and hypothyroidism were associated with CDH in clinical series. The objective of this study is to identify risk factors of chronicity in patients with headache. A total of 1,483 consecutive patients were studied. We collected information on age, gender, headache type and comorbidity. Patients were divided into three diagnostic groups: migraine and tension-type headache (CTT) diagnosis were made according to ICHD-II, and CDH fulfilling the Proposal Headache Classification for Chronic Daily Headache described by Silberstein and Lipton (in Chronic daily headache including transformed migraine, chronic tension-type headache, and medication overuse, 2001). We used descriptive statistics and Chi-square test. Our data show that age, gender and headache onset were similar in the three groups. Diabetes, hypercolesterolaemia, smoke and cardiopathy prevalence did not differ in the three groups (P > 0.05). Hypertension prevalence in CDH group (16.2%) was significantly higher than in the other two groups (migraine 7.3%; CTT 6.6%; P < 0.01). There were no differences (P > 0.05) in hypertension prevalence between CDH with and without medication overuse. CDH patients (mean age 41.8 +/- 14) referred to the Headache Center later than migraine and CTT patients (mean age 37 +/- 12) (P > 0.05). According to previous studies we found that hypertension is more frequent in CDH than in migraine and CTT. Examining this result it is possible to conclude that there exists an association between CDH and hypertension, but not that a causal relationship necessarily exists. Considering the other somatic conditions we did not find any correlation. The potential role of somatic comorbidity in CDH has to be studied in further clinical trials.

Study of Patients Self-Training Influence on Peripheral Neuropathies Diseases Diagnosis through D.I.T.A Device.

In our daily life, the sight and the sense of touch play a fundamental role in objects recognitions. This process is helped by the experience: if a subject has already seen or already touched an object in the past, he will recognize it more easily in the future. Following this assumption, the authors of this paper wanted to investigate if the experience can influence the results of a clinical examination where the subject has an active role. The attention was focused on the peripheral neuropathies diagnosis since they require an accurate assessment of several parameters including the tactile sensitivity trend. In other words, if the tests encompass an active role of the subjects, one of the main uncertainties is the self-training that influences the subject responses. This work focuses on the study of this self-training using the D.I.T.A device (Dynamic Investigation Test-rig on hAptics). Results clearly show a fundamental role of priming during "haptic modality": expert subjects, previously experienced with the tests, demonstrated better recognition of the encountered stimuli, compared to novices. Moreover, the results show that the maximum difference between the two groups of subjects is in the first part of the test. An ANOVA analysis was carried out to demonstrate that also the errors between the pins-arrays are affected by the priming.

Lewy-body dementia and responsiveness to cholinesterase inhibitors: a paradigm for heterogeneity of Alzheimer's disease?

The concept of heterogeneity of Alzheimer's disease is based on molecular, neuropathological, clinical and neuropsychological features, and also supported by the observation that Alzheimer's patients differ in their response to pharmacological interventions. Recent investigations evaluating the therapeutic potential of cholinesterase inhibitors have disclosed the existence of at least two subsets of patients with dementia, defined as 'responders' and 'nonresponders' to this therapy. In this article, Paolo Liberini and colleagues suggest that the cluster of responders to the cholinesterase inhibitors might include a significant number of subjects with a rather selective dysfunction of the cholinergic system, as in the case of Lewy-body dementia. A neuropathological demonstration of this correlation should open up new therapeutic perspectives.

Expression of nerve growth factor in brown adipose tissue: implications for thermogenesis and obesity.

The presence of nerve growth factor (NGF) and the ability of adrenergic stimulation to affect the rate of its synthesis in mouse, rat, and human brown adipose tissue (BAT) were investigated. Addition of conditioned medium, obtained from preconfluent and confluent brown adipocytes, to PC12 cells induced typical morphological changes similar to those due to NGF itself. Anti-NGF antibodies blocked this action. Moreover, NGF mRNA was detected by RT-PCR both in BAT and in brown adipocyte preparations. That NGF is synthesized in and released from brown fat cells was confirmed by immunoblotting. When the animals were exposed to low temperatures, NGF production declined. The effect of cold exposure could be mimicked by the addition of norepinephrine (NE) at day 4 or 8 (preconfluent and confluent cells, respectively). NE depletion obtained by reserpine injection induced a drastic increase of BAT NGF production. In both rat and human BAT, immunohistochemistry identified distinct anatomical structures that express the low affinity neurotropin receptor, termed p75NGFR. BAT production of NGF was higher in genetically obese rats and mice than in their lean counterparts, a difference that becomes more evident with age. Prolonged exposure to low temperature significantly decreased the BAT NGF synthesis also in obese animals. We conclude that NGF is synthesized in and released from brown fat cells, its production being inversely dependent on sympathetic activity, in both physiological and pathophysiological conditions, and increased in genetic animal models of obesity.

Characterization of dopamine receptors associated with aldosterone secretion in rat adrenal glomerulosa.

Dopamine (DA) may participate in the control of aldosterone secretion. We report that two different receptors for DA are present in rat adrenal glomerulosa: D-1, associated with stimulation of adenylate cyclase, and D-2, whose action inhibits adenylate cyclase. The adenylate cyclase system was stimulated by DA (EC50, 7.2 microM) and different DA agonists. When the D-1 receptor blocker SCH 23390 was added to the incubation medium, DA elicited a dose-dependent inhibition of adenylate cyclase (IC50, 10 microM); (-)sulpiride specifically blocked this effect. Furthermore, DA blocked angiotensin II-induced aldosterone release from glomerulosa slices in vitro. This effect was prevented by (-)sulpiride, but not by SCH 23390. The results suggest that the D-2 receptor acts to inhibit the cAMP-generating system and may be physiologically involved in the regulation of aldosterone secretion.

Long-term protective effects of human recombinant nerve growth factor and monosialoganglioside GM1 treatment on primate nucleus basalis cholinergic neurons after neocortical infarction.

Neocortical infarction induces biochemical and morphological retrograde degenerative changes in cholinergic neurons of the rat nucleus basalis magnocellularis [Sofroniew et al. (1983) Brain Res. 289, 370-374]. In the present study, this lesion model has been reproduced in the non-human primate (Cercopithecus aethiops) to investigate whether degenerative changes affecting the cortex surrounding the lesioned area and the ipsilateral basal forebrain are prevented by the early administration of recombinant human nerve growth factor alone or in combination with the monosialoganglioside GM1. Six months after surgery and treatment, the monkeys were processed either for biochemistry (choline acetyltransferase assay) or immunocytochemistry. In lesioned vehicle-treated animals, choline acetyltransferase activity significantly decreased by 28% in the cortex surrounding the injured area and by 31% in the ipsilateral nucleus basalis of Meynert when compared with values of sham-operated monkeys. These biochemical changes were fully prevented with the administration of nerve growth factor alone or in combination with the monosialoganglioside GM1. The morphometrical analysis revealed a significant shrinkage of cholinergic neurons (61 +/- 1.4% of sham-operated cell size) and loss of neuritic processes (59 +/- 10% of sham-operated values) within the intermediate nucleus basalis region of lesioned vehicle-treated animals. Although a protection of the cholinergic cell bodies within the nucleus basalis was found with both treatments, a significant recovery of the neuritic processes (84 +/- 7.2% of sham-operated values) was assessed only in the double-treated monkeys. These results indicate that the early administration of nerve growth factor alone or in combination with the monosialoganglioside GM1 induces a long-term protective effect on the nucleus basalis cholinergic neurons in cortical injured non-human primates.

Differential expression of fetal and mature tau isoforms in primary cultures of rat cerebellar granule cells during differentiation in vitro.

The molecular mechanism(s) responsible for the differential expression of various tau protein isoforms as well as their functional role in morphogenesis, neurofibrillary tangle formation and neurodegeneration have not been completely clarified. We found that the expression of tau proteins in primary cultures of cerebellar granule cells from neonatal rat brain is a developmentally regulated process affecting tau synthesis at different levels. Changes in tau RNA splicing are clearly demonstrated by PCR data showing the switching on of the mRNA containing four internal repeats by DIV 6 and the switching off of the mRNA containing three internal repeats after DIV 12. The changes in mRNA levels of the different tau isoforms during development in vitro occur in parallel with changes in tau protein expression, both qualitatively and quantitatively, as shown by Western analysis of protein extracts from granule cells at different DIV with an anti-tau polyclonal antibody. Finally, as indicated by MAP2 and tau immunocytochemistry data, the switch in tau protein expression appears to be contemporary with neurite outgrowth and cell differentiation. Our data suggest that a differential expression of various tau proteins parallels the degree of cell maturation.

Encapsulated genetically engineered fibroblasts: release of nerve growth factor and effects in vivo on recovery of cholinergic markers after devascularizing cortical lesions.

Genetically engineered rat fibroblasts producing nerve growth factor (NGF) were encapsulated in alginate-polylysine-alginate gels with the objective to produce viable "minifactories" continuously producing and secreting NGF into the rat brain. Microencapsulated fibroblasts (NGF secretors and NGF non-secretors) were placed onto the surface of the lesioned rat cortex (unilateral devascularizing lesion) and animals were sacrificed 30 days after surgery. Fibroblasts NGF-non secreters normally produce tumors after implantation, therefore, they were irradiated prior to encapsulation. Three other experimental groups were studied in parallel: non-lesioned (controls), lesioned rats receiving "empty" alginate spheres and lesioned animals without treatment and microspheres. Biochemical analysis of microdissected brain tissues of lesioned animals treated with encapsulated NGF-secretor fibroblasts showed a significant increase in choline acetyltransferase (ChAT) activity in cortices adjacent to the lesion but not far from it (entorhinal cortex). This may indicate a gradient of concentration of the released NGF and/or differential responsivity of lesioned vs non-lesioned target tissue. ChAT enzymatic activity in the microdissected nucleus basalis magnocellularis (NBM) was significantly decreased (P < 0.05) in all lesioned animals treated with spheres without fibroblasts and those with fibroblasts not secreting NGF. Morphometric analysis of ChAT-IR and low affinity NGF-receptor IR cholinergic neurons in the middle portion of the NBM shows a prevention of neuronal shrinkage and extensive neuropil in animals treated with microencapsulated NGF-secretor fibroblasts. The results of this study demonstrate that NGF from encapsulated genetically engineered fibroblasts can be secreted for at least long enough to prevent degenerative changes of cholinergic neurons in the NBM.

Differential effect of acute reserpine administration on D-1 and D-2 dopaminergic receptor density and function in rat striatum.

In the present study we investigated the effects of a single injection of reserpine on dopamine receptor function in rat striatum. The measurement of adenylate cyclase activity indicated that cAMP formation induced by the selective D-1 agonist SKF 82526 was greatly increased 2 and 20 h after reserpine; in contrast, the selective D-2 agonist bromocriptine inhibited by the same extent and with the same potency AC in controls and reserpinized striata. In addition, the density of both D-1 and D-2 receptors, as measured in binding studies with the selective D-1 antagonist [(3)H]SCH 23390 and with [(3)H]spiroperidol respectively, was not affected by the treatment. These results suggest that acute reserpine administration induces a selective up-regulation of the transduction mechanisms associated with D-1 receptors, without changing the functional activity of D-2 receptors.

Nerve growth factor treatment restores [3H]QNB binding site density in adult rat subjected to cortical infarction.

The effects of unilateral cortical devascularization and nerve growth factor treatment on muscarinic cholinoceptors labelled with [3H]QNB were investigated in rat cerebral cortex. The lesion induced a significant ipsilateral decrease in the density (Bmax) but not in the affinity (kD) of [3H]QNB binding sites. Intraventricular infusion of 2.5 S mouse nerve growth factor (NGF) increased the density of binding sites such that Bmax values of lesioned animals treated with NGF were not different from control subjects.

Tau protein immunolocalization in fetal and adult human spinal cord.

In the present study, the monoclonal antibody Alz-50 has been used to determine and compare the immunohistochemical localization of phosphorylated tau proteins in the developing and normal adult spinal cord. At all stages of fetal life Alz-50 fiber immunoreactivity was observed in the dorsal roots, in the dorsal and dorsolateral funiculi, and in restricted regions of the dorsal horn. Alz-50 immunoreactivity was also demonstrated in the dorsal root ganglion neurons. In the adult spinal cord a consistent pattern of Alz-50 fiber immunoreactivity was localized in the superficial layers of the dorsal horn (lamina I and II) but not in dorsal and dorsolateral funiculi and in the dorsal root ganglion. Comparable results in fetal specimens have been obtained employing PHF-1, a monoclonal antibody generated against paired helical filament proteins from Alzheimer brains, while no significant immunostaining for PHF-1 was observed in the adult spinal cord. In addition, the staining with monoclonal and polyclonal anti-tau antibodies overlapped with that of Alz-50. The transient, selective pattern of Alz-50 and PHF-1 immunoreactivity may disclose some relevant functions of tau proteins during somatosensory pathway development.

NGF-mediated synaptic sprouting in the cerebral cortex of lesioned primate brain.

In the present study, coronal brain sections of cortically devascularized non-human primates (Cercopithecus aethiops) were used to assess the lesion-associated synaptic loss, and the effect of exogenous nerve growth factor (NGF) in preventing or reversing this neurodegeneration. The sections were immunolabeled with antibodies against the synaptic marker protein synaptophysin (SYN), as well as choline acetyltransferase (ChAT) and parvalbumin (PV) markers that identify cholinergic neurons and interneurons, respectively. We found that, compared to sham-operated animals, in the lesioned vehicle treated animals SYN immunoreactivity near the lesioned site in the frontoparietal cortex was decreased by 31%. Similarly, corrected optical density values of immunostained sections specific for ChAT in the nucleus basalis of Meynert (ipsilateral to the lesion) decreased by 20% and PV-immunoreactive neurons near the lesion decreased by 47%. In contrast, NGF-treated lesioned animals showed levels of SYN, ChAT, and PV immunoreactivity similar to sham controls. These results are consistent with previous studies and support the view that NGF may not only prevent neurodegenerative changes after neocortical infarction by protecting vulnerable neurons, but also is capable of inducing sprouting and synaptogenesis.

Neocortical infarction in subhuman primates leads to restricted morphological damage of the cholinergic neurons in the nucleus basalis of Meynert.

The aim of the present study was to investigate the long-term effect of cortical infarction on the subhuman primate (Cercopithecus aethiops) basal forebrain. The lesion, carried out by cauterizing the pial blood vessels supplying the left fronto-parieto-temporal neocortex, induced retrograde degenerative processes within the ipsilateral nucleus basalis of Meynert. The morphometrical analysis revealed that significant shrinkage of cholinergic neurons and loss of neuritic processes were localized within the intermediate regions of the nucleus basalis. The average cross-sectional areas of choline acetyltransferase-immunoreactive neurons in the intermedio-ventral (Ch4iv) and intermedio-dorsal (Ch4id) nucleus basalis were decreased to 62.5 +/- 9.5 and 58.0 +/- 8.6%, respectively, of the sham-operated values. Although an apparent loss of Nissl-stained magnocellular neurons in Ch4iv and Ch4id was found by applying a quantitative analysis based on a perikaryal-size criterion, data obtained by the quantification of immunostained material failed to reveal any significant decrease of cholinergic cell density. Results are discussed in view of future application of this ischemic model to study processes of retrograde degeneration following cortical target removal and to assess potential neurotrophic and neuroprotective properties of pharmacologic agents.

Repeated reserpine administration up-regulates the transduction mechanisms of D1 receptors without changing the density of [3H]SCH 23390 binding.

Behavioural studies have shown that stimulation of D1 receptors, which is uneffective in normal rats, induced strong hypermotility in rats pretreated with reserpine for 5 days. On this basis, we investigated D1 receptor plasticity using the 5-day treatment with reserpine (1 mg/kg; s.c.) as an experimental model. The function of striatal D1 receptors was determined both in binding studies with [3H]SCH 23390 and by measuring formation of cAMP in response to the selective agonist, SKF 82526. The results indicate that the responsiveness of adenylate cyclase (AC) to D1 receptor stimulation was markedly increased after reserpine administration, while no significant changes were found in [3H]SCH 23390 binding site density. Moreover, formation of cAMP after stimulation of Gs protein with GppNHp was markedly enhanced in dopamine (DA)-depleted rats; the responsiveness of AC to forskolin, which directly stimulates the AC catalytic unit, was not affected by reserpine administration. These data indicate that reserpine-induced D1 receptor up-regulation is apparently mediated by a marked enhancement of the coupling efficiency of Gs protein, suggesting that the D1 behavioral supersensitivity does not correlate with the density of D1 receptors, but is reflected by a selective up-regulation of their transduction mechanisms.

Olfaction in Parkinson's disease: methods of assessment and clinical relevance.

Several neurological conditions have been reported to be associated with peripheral or central deficits of olfactory system. In recent years particular emphasis has been placed on the early and severe olfactory impairment in Parkinson's disease (PD), in which limited neuropathological studies have revealed a marked dopaminergic deficit in the olfactory tubercles. Moreover, indirect evidence suggests that dysfunction of the dopaminergic pathways from mesencephalon to the piriform cortex may play a role in olfactory impairment in PD. A large number of clinical studies have reported that olfactory loss in idiopathic PD is bilateral, present in hemiparkinsonism, unrelated to the stage or clinical subtype of the disease, and independent of antiparkinsonian medication. In addition, major olfactory alterations have been reported in familial PD and dementia with Lewy bodies but not in progressive supranuclear palsy and essential tremor. These findings might stimulate further research targeted to determine the biological substrate of dissimilar olfactory performances in these movement disorders. The present review summarizes standardized procedures for the assessment of olfactory acuity (detection threshold), identification (multiple choice odor naming), discrimination (differentiation between similar/dissimilar odorants), and memory (recognition of a substance previously smelled). Specific suggestions concerning the psychometric and neuropsychological evaluation of PD patients are provided.

Alz-50 immunoreactivity in the central nervous system of adult rat and primate.

The purpose of this work was to investigate the distribution and density of Alz-50 immunoreactivity in the central nervous system of normal adult and cortically injured rats and primates (Cercopithecus aethiops). In control animals of both species a consistent pattern of fiber immunoreactivity was detected within the hypothalamus (arcuate nucleus and median eminence) and the spinal cord (posterior horn and dorsal root nerve). Immunoreactive perikarya were predominantly observed throughout the anterior region of the third ventricle. An identical localization and density of Alz-50 staining was observed in lesioned animals. These experiments reveal that the pattern of Alz-50 immunoreactivity is not affected by the neurodegenerative processes that follow the cortical devascularizing lesion. These observations suggest that the monoclonal antibody Alz-50, besides recognizing cytoskeletal components in degenerating neurons, reacts with specific epitopes located in the hypothalamus and spinal cord of normal mammalian central nervous system.

Acute quadriplegia with delayed onset and rapid recovery. Case report.

The authors describe a patient with severe head injury and sepsis who became acutely quadriplegic 3 days postinjury because of a critical illness polyneuropathy (CIP) and critical illness myopathy (CIM), which resolved rapidly after treatment of the underlying infection. In only 3 days the patient developed septic shock together with flaccid quadriplegia and absent deep tendon reflexes with no clinical or radiological evidence of central nervous system deterioration. Neurophysiological studies showed an acute axonal sensorimotor polyneuropathy, whereas the clinical course strongly suggested a concurrent myopathy. A severe Staphylococcus epidermidis infection accompanied by bacteremia was treated and the patient recovered fully within a few days. Although the case described here is unique because of its very early onset and rapid resolution, CIP and CIM are frequent complications of sepsis and multiple organ failure. The authors suggest that severely head injured patients with sepsis should be evaluated for CIP and CIM when presenting with unexplained muscle weakness or paralysis.

Identification of D-2 dopaminergic receptors in bovine adrenal cortex.

Dopamine receptors in bovine adrenal cortex have been studied by using 3H-(-)sulpiride as selective ligand. The specific binding is saturable and the Scatchard analysis reveals a single component with a Kd of 6.2 nM and a Bmax of 8 fmoles/mg protein. The characterization indicates that the binding is rapid, reversible, stereospecific, Na+ - and temperature- dependent. Moreover its pharmacological profile is superimposable to that of D-2 receptors in the striatum, thus suggesting that central and peripheral D-2 receptors are identical.