Correlation between VITOM(®) videocolposcopy and histopathology for pathognomonic grading criteria.

OBJECTIVE: "Every colposcopic criterion must be mirrored by histopathology". We investigated the histomorphologic equivalent of four colposcopic criteria, which are associated with CIN 2 and/or CIN 3 and therefore called pathognomonic. PATIENTS AND METHODS: We diagnosed inner border sign, ridge sign, rag sign and/or cuffed gland openings using VITOM(®) videocolposcopy in 255 patients which are consistent with major change. Histopathologic examination included immunohistochemical staining for p16, Ki 67 and stathmin-1 and micro-photographic documentation. RESULTS: The histopathologic pattern specific for each of the four pathognomonic colposcopic criteria was reproducibly identified: inner border sign showed a sharp demarcation between low- and high-grade CIN, in ridge sign high-grade CIN adjoined directly the squamocolumnar junction, in rag sign, high-grade CIN was detached from stroma, and in cuffed gland openings, the entrance to a gland was rimmed by CIN, respectively. In 255 patients, the leading pathognomonic sign was inner border in 12.1 %, ridge in 34.1 %, rag in 18 %, and cuffed glands in 35.7 %, respectively. Inner border sign, ridge sign, rag sign and/or cuffed gland openings were associated with CIN 2 or 3 in 97, 98, 98 and 98 %, respectively. In 153 out of 255 patients, we found a combination of pathognomonic signs with ridge sign being the most frequent combined criterion (in 21 % of patients as second pathognomonic sign). CONCLUSION: The morphology of the four pathognomonic colposcopic criteria, inner border sign, ridge sign, rag sign and cuffed crypt openings, is reproduced in histopathology. These criteria are highly associated with CIN 2 or CIN 3.

Study investigating pharmacokinetic interaction between theophylline and roflumilast in healthy adults.

OBJECTIVE: To investigate whether a pharmacokinetic drug-drug interaction exists between theophylline (THEO), a CYP1A2 substrate with a narrow therapeutic index, and the concomitant substrate roflumilast (ROF), a novel selective PDE4 inhibitor partially metabolized by CYP1A2. MATERIALS AND METHODS: In an open-label, 2-period, crossover study, Treatment A (oral ROF 500 µg q.d. on Days 6 - 10 in addition to oral THEO 375 mg b.i.d. on Days 1 - 10) and treatment B (oral ROF 500 µg q.d. on Days 1 - 5) were administered consecutively in random order to each of 24 healthy adult subjects. Both periods were separated by a wash-out phase of at least 10 days. Plasma samples for pharmacokinetic evaluation (AUC, Cmax, t1/2, tmax) including percent peak-trough fluctuation (%PTF) of THEO were taken. Point estimates and the 90% confidence interval of the geometric mean ratios were calculated for AUC and Cmax and descriptive statistics for other pharmacokinetic parameters. RESULTS: Concomitant administration of ROF did not alter pharmacokinetics of THEO. With coadministered THEO, only steady-state total exposure to ROF (AUC) was increased by 28% whereas other pharmacokinetic parameters (t1/2, Cmax, tmax) of ROF and of the active metabolite roflumilast-N-oxide (R-NO), its main contributor to the pharmacodynamic effects, remained unchanged. CONCLUSIONS: Neither ROF nor its main metabolite had any impact on the metabolism of the concomitant CYP1A2 substrate THEO in humans. Though co-administration of THEO resulted in a minor increase (28%) in total ROF exposure, no safety or tolerability concerns and no altered total PDE4 inhibition of both ROF and R-NO, were observed.

Cervical Strip Biopsy for High-Grade Cervical Intraepithelial Lesions: a Valid Alternative to Conventional Punch Technique.

Introduction: To evaluate, if targeted strip biopsies decrease trauma/pain perception while maintaining diagnostic accuracy in patients with the diagnosis of high-grade squamous intraepithelial lesions of the uterine cervix. Patients and Methods: Between July 1st and December 31st 2014 we performed colposcopically directed strip biopsies in 102 patients with colposcopic suspicion of high-grade squamous intraepithelial lesions of the uterine cervix. We used a 3 mm curette for harvesting tissue samples under VITOM® videocolposcopy. So far, 60 patients underwent additional loop excision. Histologic examination of strip biopsies and loop specimens included routine hematoxylin and eosin staining as well as immunohistochemical staining for p16, Ki 67 and stathmin-1. Results: 55 patients (53 %), were histologically diagnosed with cervical intraepithelial neoplasia grade 3 on strip biopsies. Adenocarcinoma in situ was diagnosed in 2 patients (2 %), cervical intraepithelial neoplasia grade 2 in 35 patients (34 %), and cervical intraepithelial neoplasia grade 1 in 10 patients (10 %). The agreement between histologic results of strip biopsy and loop specimen was highly significant: In all 60 strip biopsies diagnosed with high-grade squamous intraepithelial lesions this diagnosis was confirmed histologically during follow-up loop specimen excision (high-grade squamous intraepithelial lesions in 58 patients, invasive disease in 2 patients). The pain level experienced during strip biopsy was rated on average 0.25 on a scale from 0 to 10. No clinically significant bleeding was reported. Conclusion: Targeted strip biopsies with a 3 mm curette are a reliable procedure to diagnose high-grade squamous intraepithelial lesions of the uterine cervix and yield high patient satisfaction (Video 1).

Cholinergic effects on spike-density and burst-duration of medullary respiration-related neurones in the rabbit: an iontophoretic study.

Cholinoceptive properties of 180 medullary respiration-related neurones (RRN) were studied in urethane-anaesthetized rabbits. Acetylcholine (ACh) and agonists, as well as antagonists of muscarinic receptors and nicotinic receptors, were administered iontophoretically. Respiration-related neurons were classified with respect to the correlation of their activity with the activity of the phrenic nerve: phase-bound inspiratory (I) and expiratory (E) neurones and phase-spanning expiratory-inspiratory and inspiratory-expiratory neurones were discriminated. Acetylcholine altered the activity of 170 respiration-related neurones. In 49 cells the discharge-rate (f) alone was affected. In 44 respiration-related neurones the burst-duration (tbd) only was altered. In 77 respiration-related neurones the discharge rate and burst duration were affected simultaneously. Inhibitory actions of ACh prevailed over excitatory actions on inspiratory neurones. In expiratory neurones the discharge rate was increased or decreased in about the same number of cases. In most phase-spanning neurones the discharge rate was increased. Cholinergic inhibition of burst duration was observed in most respiration-related neurones. In all phase-types of respiration-related neurones muscarinic as well as nicotinic actions of ACh were demonstrated. In inspiratory and expiratory-inspiratory neurones muscarinic effects on the discharge rate prevailed over nicotinic effects. More nicotinic than muscarinic effects on discharge rate were observed in expiratory and inspiratory-expiratory neurones. Cholinergic effects on burst duration in about the same number of respiration-related neurones were mediated by muscarinic or by nicotinic receptors, respectively. Various types of cholinoceptors may be involved in these effects. The results suggest that cholinergic mechanisms play an important role in the control of the central regulation of respiratory movements. The functional significance of cholinergic effects on respiration-related neurones is discussed with special emphasis of effects on burst duration.

Immuno- and histochemical studies on galactan and human blood group-related receptors in the bovine lung.

Using a monoclonal anti-galactan antibody and streptococcus B type II antibody, the distribution of lung galactan could be demonstrated for the first time in a vertebrate organ. In addition to the immunochemical demonstration of the bovine lung galactan, a human blood group A-like glycoprotein is detectable by lectinological methods in the bovine lung tissue. Various other lectin-receptors, for instance those of the peanut lectin (PNA) or for lectins with L-fucose (UEA) and N-acetyl-lactosamine (ECA) specificity show a typical staining pattern in bovine lung.

A stepper motor-driven microelectrode positioner.

The mechanical elements and the electronic control system from a stepper motor-driven microelectrode positioner is described. The unit embodies a high-precision small step angle hybrid motor. The compact, rugged and totally concentric design of the mechanic, by a spindle mechanism achieves the necessary precision by translating the stepwise rotations of the motor into steps of linear movement. The system takes advantage of commercially available low friction parts such as ball bearings, ball bushings and axles with hardened surfaces. The related electronic control unit is designed around the most recent integrated circuitry which is both sophisticated and economical. Though the described system is designed to be built in an average departmental workshop it compares favorably with more expensive commercial units and in some aspects outperforms them.

A device for spike train sampling with built-in memory.

The described interface to a digital computer measures interspike interval durations with a resolution of 10 microseconds. A built-in first-in first-out (FIFO) memory relieves the host computer from frequent I/O intensive tasks. The internal FIFO buffer can store up to 512 data words (wordlength is 16 bit) and works on the dual-port principle. This way the acquisition of a neuronal spike train is completely independent of the computer's simultaneously ongoing data access. A simple handshake protocol between the interface and the computer prevents any overhead communication. The buffer architecture of the instrument releases the host computer from high speed I/O handling schemes like real-time, clock-controlled, polling or interrupt procedures, that would request assembly language support. The body of two software, driver routines in the BASIC and the PASCAL language is presented. A complete and detailed schematic diagram of the circuitry is included.

An electronic device for accurate quantification of neuronal mass activity based on a digital integration method.

Quantification of different kinds of neuronal mass activity was performed using an integrative method. For this purpose an electronic device was designed and constructed which is based on a fully digital approach. High precision of the method and an easy way to interface with a digital computer are the advantages of this strategy when compared to other techniques. Performance of quantification and the possibility of the universal use of the device are demonstrated by two examples. (1) Quantitative analysis of whole nerve bundle activity is demonstrated in the case of phrenic nerve mass activity. (2) The usefulness of the digital integrator is further verified in quantification of theta waves of the human EEG as a rather different field of application. In addition, the device is valuable as a tool for the analysis of EMGs. The described digital integrator has many advantages compared to conventional analog integrators, such as autorange feature, very high dynamic range, excellent long-term stability, in principle unlimited integration times and interface capability to a digital computer; it avoids all problems associated with the application of high-impedance integrator operational amplifiers.

Involvement of fast synaptic inhibition in the generation of high-frequency oscillation in central respiratory system.

During moderate hypercapnia, spectrum analysis of efferent phrenic nerve activity (PNA) of urethane anesthetized rabbits revealed high-frequency oscillations (HFO). The spectral peak (mean frequency 111 Hz) was reversibly eliminated by the glycine receptor antagonist strychnine administered into the 4th cerebral ventricle. In contrast, blockade of brainstem GABAA receptors by bicuculline did not abolish HFO but even reinforced or induced HFO. Thus, there is evidence that fast synaptic inhibition via glycine receptors is involved in the generation of HFO within the medullary respiratory center, whereas GABA may play a modulatory role.

Effects of excitatory amino acids and neuropeptide Y on the discharge activity of suprachiasmatic neurons in rat brain slices.

Effects of L-glutamate, AMPA, NMDA and NPY on the discharge activity of neurons located in the ventral subdivision of the suprachiasmatic nucleus were examined in submerged coronal slices of the rat hypothalamus. All substances were bath applied. Application of L-glutamate (14 neurons examined) induced an excitatory response in 8 suprachiasmatic neurons (+248.9 +/- 122.24%, mean +/- S.E.M.; P < 0.001). A biphasic response, i.e. an initial transient excitation (+54.3 +/- 8.21%; P < 0.001) succeeded by an inhibition (-66.2 +/- 9.31%; P < 0.001), was observed in 6 neurons. Application of AMPA (36 neurons examined) resulted in an excitation of 31 neurons (+209.2 +/- 58.58%; P < 0.0001). Application of NMDA (57 neurons examined) induced an excitation in 34 neurons (+253.8 +/- 91.18%; P < 0.0001), but an inhibition in 8 neurons (-757 +/- 6.52; P < 0.0001). Biphasic effects of NMDA with an excitatory component (+58.7 +/- 9.94%; P < 0.0001) succeeded by an inhibitory component (-62.0 +/- 8.07%; P < 0.0001) were observed in 13 neurons. In 5 of 13 examined cases, the inhibitory component of neuronal responses to NMDA was significantly attenuated by the simultaneous application of strychnine (attenuation was 56%; P < 0.05). The application of NPY (40 neurons examined) induced significant effects on the discharge rate of 29 suprachiasmatic neurons. 18 of these neurons were inhibited (-59.3 +/- 6.39%; P < 0.0001) whereas 11 neurons were excited (+156.6 +/- 107.222%; P < 0.001) by NPY. In 8 of 11 neurons examined, the NPY-induced inhibition was significantly attenuated by 92% during simultaneous application of strychnine (P < 0.001). In 23 NPY-sensitive neurons, the discharge activity was also affected by NMDA. Neurons excited by NPY were also excited by NMDA (8 cells). In neurons inhibited by NPY, application of NMDA induced either an inhibition (3 cells) an excitation (5 cells) or a biphasic effect (7 cells). Results suggest a direct excitatory effect of AMPA, NMDA and NPY on suprachiasmatic neurons. In contrast, inhibitory actions of NMDA and NPY are considered induced by an activation of inhibitory interneurons. Antagonistic effects of strychnine suggest an involvement of glycinergic interneurons in a subpopulation of neurons inhibited by NMDA and in most neurons inhibited by NPY. The involvement of inhibitory mechanisms in photic entrainment of the circadian system is discussed. An integrative model of excitatory and inhibitory actions of EAA and NPY on suprachiasmatic neurons is proposed.

Concurrent fast and slow synchronized efferent phrenic activities in time and frequency domain.

In urethane-anesthetized or decerebrated vagotomized rabbits efferent multifiber activity of the phrenic nerve was investigated for synchronized activities both in time and frequency domains. When respiratory drive was steadily increased by either an elevation of end-tidal CO2 concentration or i.v. administration of 4-aminopyridine, medium-frequency oscillations (MFO) first increased, then decreased and finally became absent. The power of high-frequency oscillations (HFO) steadily rose with increasing respiratory drive. In contrast to HFO which revealed a unimodal spectral peak of mostly small bandwidth, the MFO spectrum in most cases consisted of a broad complex. This complex in some cases was composed of two distinct peaks, i.e. MFO were heterogenous. The low- and high-frequency fractions of the MFO complex were related predominantly to the first and last third of inspiration, respectively. Examination of the on-going multifiber activity of the phrenic nerve with an expanded time scale revealed that lower frequency MFO probably result from synchronized ramp-like wave activity during early and mid-inspiration. The duration of the observed ramps well matched the corresponding MFO frequency. We suggest that these ramps might result from propagated synchronized waves of high-threshold phrenic motoneurons. During the last part of inspiration, however, MFO, like HFO, resulted from burst-like synchronized discharge of phenic motoneurons. Thus HFO are superimposed on ramp-like and burst-like activity of the MFO. It is assumed that the decline of MFO at high respiratory drive may be due to the increasing strength of HFO bursts which interrupt ramp activity in the MFO range and thus let MFO appear 'invisible' to the recording electrode. Both MFO and HFO were visually detectable in postinspiration.

Effects of corticotropin-releasing factor on central respiratory activity.

Effects of corticotropin-releasing factor (CRF) on central regulation of respiration were studied in urethane-anaesthetized, vagotomized, paralyzed and artificially ventilated rabbits. Injections of CRF into the IVth cerebral ventrile (i.c.v.) resulted in an increase of the neuronal tidal volume (nVt), but had only minor effects on the duration of respiratory phases. Microinjection of CRF into the ventral parabrachial region of the pons (pneumotaxic center) resulted in a reduction of nVt and of the respiration rate. Respiratory effects of CRF administered i.c.v. or into the pontine parabrachial region were antagonized by alpha-helical-CRF, indicating a receptor-mediated action of CRF. Respiratory effects of CRF were not significantly affected after blocking ganglionic transmission with chlorisondamine. Neither mean arterial blood pressure nor heart rate were significantly affected by the injection of CRF into the IVth ventricle or the rostral pons. Results suggest an involvement of CRF in the central regulation of respiratory movements. CRF may be involved in the adaptation of respiration to stress.

Dopaminergic control of respiration as shown by effects of 4-aminopyridine.

The dopaminergic control of respiration in conscious and urethane-anaesthetized rabbits, was studied by comparing the respiratory effects of 4-aminopyridine alone (4-AP; 1 mg/kg i.v.) and those after the administration of dopamine antagonists (domperidone and haloperidol; 1 mg/kg each). The respiratory rate in conscious rabbits was increased by 4-AP. After domperidone this increase was reduced and preceded by a transient decrease. In spontaneously breathing, anesthetized rabbits there was a transient reduction after which the respiratory rate was increased by 4-AP; tidal volume was affected in an inverse manner. After domperidone, the excitatory effect of 4-AP on respiratory rate and the inhibitory effect on tidal volume were blocked. The effects of 4-AP on respiratory rate were prevented by vagotomy. In anesthetized, vagotomized, paralyzed and artificially ventilated rabbits (VPV animals) the peak amplitude of the integrated phrenic nerve activity ("phrenic activity') was increased by 4-AP. After pretreatment with haloperidol this effect of 4-AP on phrenic activity was reduced while the respiratory rate was now increased. In VPV animals with denervated carotid bodies the excitatory effect of 4-AP on phrenic activity was strongly enhanced and respiratory rate was increased. These effects were slightly reduced but not blocked by haloperidol. It is concluded that endogenous dopamine is involved in the control of respiration through effects on peripheral mechanisms (inhibition of inspiratory activity and enhancement of respiratory rate) as well as on central mechanisms (stimulation of inspiratory activity and reduction of respiratory rate).

GABAB receptor mediated effects on central respiratory system and their antagonism by phaclofen.

The role of GABAB receptors in control of central respiratory system was evaluated by cycle-triggered averaging of phrenic nerve activity (PNA) of the rabbit. Blockade of GABAB receptors of the caudal brainstem by intracerebroventricular administration of phaclofen augmented PNA, decreased the duration of inspiration and to about the same extent increased the duration of expiration thus unmasking intrinsically active GABA. Analogously, stimulation of brainstem GABAB receptors by exogenous baclofen decreased PNA. Preceding administration of larger doses of phaclofen could block the effects of baclofen. It is proposed that GABAB receptors are involved in tonic and phasic modulation of central respiratory activity.

Evidence for an involvement of NMDA and non-NMDA receptors in synaptic excitation of phrenic motoneurons in the rabbit.

The action of endogenous excitatory amino acids on phrenic motoneurons was studied in anesthetized, vagotomized, paralyzed and artificially ventilated rabbits. The NMDA receptor antagonists APV and ketamine, as well as the non-NMDA receptor antagonists GAMS and DNQX were administered by microinjection into the ventral horn of the spinal segments C3-C5. Injection of each antagonist resulted in a reversible reduction of the phrenic nerve activity. Results suggest an important function of endogenous excitatory amino acids in the excitation of phrenic motoneurons. NMDA as well as non-NMDA receptors are involved. The functional role of both receptor types in bulbospinal neurotransmission is discussed.

Serotonergic control of phrenic motoneuronal activity at the level of the spinal cord of the rabbit.

The role of serotonin (5-hydroxytryptamine, 5-HT)-mediated modulation of phrenic motoneuronal activity was evaluated by microapplication of 5-HT and methysergide into the phrenic nuclei of the rabbit. 5-HT facilitated phrenic nerve activity (PNA) considerably resulting in a long-lasting augmentation of the peak amplitude of integrated PNA. In contrast, the blockade of intrinsically active 5-HT by methysergide decreased PNA and led to a strong reduction of the peak amplitude of integrated PNA. MDL 72222 was ineffective. Blockade of 5-HT receptors by preceding administration of methysergide effectively abolished the effects of microinjected 5-HT. Respiratory timing was unaffected by both the agonist and the antagonist. These results suggest that a considerable portion of the facilitatory influence of caudal raphe nuclei on central respiratory activity takes place at the phrenic nuclei level.

Axonal projections and primary afferents of bulbar respiratory modulated and unmodulated neurons travelling with vagal nerve or spinal cord in the rabbit.

Respiratory modulated neurons of the rabbit were examined for efferent projections and afferent connections via the spinal cord or the vagal nerves. Variability of latency of responses proved to be related to latencies. When latency of evoked potentials is plotted versus variability of latency, monosynaptic responses can apparently be discriminated from oligo- or polysynaptic responses. Latency of antidromic responses depends from stimulus frequencies used. Results showed that only 12% of all respiratory modulated neurons have spinal descending axons conducting at 9-45 m per sec and 8% of the inspiratory group of neurons possess axons running along with the vagal nerves conducting at 20-37 m per sec. It is concluded that the respiratory network in the rabbit is essentially different from that in the cat.

Microelectrophoretic application of metabolic modifiers of the phosphofructokinase-hexosediphosphatase-system onto various types of bulbar respiratory neurons.

Three metabolic modifiers of the PFK-HDPase system were applied to bulbar respiratory neurons and, for comparison, also to unspecific cells. F-6-P did not alter the spike density of the majority of the respiratory and about half of the unspecific neurons. When responding, IE units were activated and unspecific cells were inhibited. Citrate did not alter the spike density of about half of all neurons tested. When responding, excitation prevailed in IE and E units, while unspecific cells were inhibited. AMP did not alter the spike density of the majority of the respiratory and about half of the unspecific cells. When responding, I neurons were activated, inhibition preponderated in IE units and both effects occurred in unspecific cells. The large number of zero effects suggests that the activity of only part of the cells is governed by the PFK-HDPase system.

Microelectrophoretic application of antagonists of putative neurotransmitters onto various types of bulbar respiratory neurons.

Seven antagonists of putative neurotransmitters were applied to bulbar respiratory neurons and, for comparison, also to unspecific cells. The antagonists exerted distinct effects when released alone, permitting to draw conclusions about receptor properties of the various cell types. With strychnine, specific antagonist of glycine, excitation prevailed in EI, I and E neurons. With bicuculline, specific antagonist of GABA, excitation preponderated in EI and E cells. About half of the unspecific neurons were activated and the remainder were unresponsive. GDEE (glutamatediethylester), antagonist of glutamate, excited part of the IE neurons and inhibited part of the E units, while the remainder of both types as well as 2 EI cells tested were not affected. With flupentixol, antagonist of dopamine, excitation prevailed in I neurons. About half of the IE and E units remained unaffected, while in the remainder E cells inhibition preponderated over excitation. With yohimbine, an alpha-adrenoceptor blocker, inhibition prevailed in E units. The two EI as well as the majority of the I neurons remained unaffected, with two cells of the latter type being activated. Propranolol, a beta-adrenoceptor blocker, inhibited about half of the E neurons, while the remainder as well as most IE and the 2 EI cells tested were not affected. Cyproheptadine, an antagonist of 5-HT, excited most E neurons. As concerns NE-receptors, those of the alpha-type might be involved in activation of part of the E cells only, whereas all other NE effects (inhibition or activation) are mediated by CNS-specific receptors different from the alpha- and beta-type. 5-HT effects apparently are mediated by two different receptor types.

Inhibitory action of vagal nerve and spinal cord afferents on discharge pattern of bulbar respiratory modulated neurons.

Single vagal nerve stimuli delivered during burst discharge of respiratory modulated neurons entailed lengthening of an interspike interval in 37% of all units tested. Occasionally stimuli delivered to the spinal cord caused lengthening of interspike intervals in respiratory modulated neurons and in other bulbar units the discharge of which was not modulated with respiration. Vagal stimuli often entailed lengthening of the following interspike interval, while the interval during which stimuli were applied remained unaltered; latencies suggested that inhibitory pathways were polysynaptic. In expiratory-inspiratory (EI) and inspiratory (I) neurons, lengthenings of intervals roughly doubled those noted in inspiratory-expiratory (IE) and expiratory (E) units. Lengthening of intervals depended on stimulus incidence within the intervals and was greatest when stimuli were delivered during the late part of the intervals. In EI and I units, lengthening of intervals occurred mainly during the middle part of burst discharge, in IE and E neurons, however, towards the end of the burst. Conditioning of discharge of respiratory modulated neurons could be demonstrated.