Site Of Drug Application Research Articles

Estimation of skin absorption of chemicals with Fick's diffusion model

The skin is not only the site of drug application, but also the site of exposure to various chemical substances. Prediction of skin permeability, body absorption, and local skin concentration of chemicals are very important to assure efficacy and safety. Most in silico models for skin permeation of chemical are based on the prediction of its permeability coefficient (P, unit cm/s), and a certain level of prediction accuracy has been obtained. On the other hand, the amount absorbed in the body of a chemical is calculated from the sum of the amount of skin permeation and the amount under the stratum corneum (viable epidermis and dermis). The amount of skin permeation can be calculated from the P value as mentioned above, however, the amount under the stratum corneum, which is related to the local skin safety of the chemical, cannot be predicted. This article describes the principles of permeation of chemicals across the skin, the relationship between skin permeability and concentration in the skin, and the method for calculating the chemical concentration under the stratum corneum using skin permeation parameters.

Postsynaptic activity of inhibitory neurons evokes hemodynamic fMRI responses

Functional MRI responses are localized to the synaptic sites of evoked inhibitory neurons, but it is unknown whether, or by what mechanisms, these neurons initiate functional hyperemia. Here, the neuronal origins of these hemodynamic responses were investigated by fMRI or local field potential and blood flow measurements during topical application of pharmacological agents when GABAergic granule cells in the rat olfactory bulb were synaptically targeted. First, to examine if postsynaptic activation of these inhibitory neurons was required for neurovascular coupling, we applied an NMDA receptor antagonist during cerebral blood volume-weighted fMRI acquisition and found that responses below the drug application site (up to ~1.5 mm) significantly decreased within ~30 min. Similarly, large decreases in granule cell postsynaptic activities and blood flow responses were observed when AMPA or NMDA receptor antagonists were applied. Second, inhibition of nitric oxide synthase preferentially decreased the initial, fast component of the blood flow response, while inhibitors of astrocyte-specific glutamate transporters and vasoactive intestinal peptide receptors did not decrease blood flow responses. Third, inhibition of GABA release with a presynaptic GABAB receptor agonist caused less reduction of neuronal and blood flow responses compared to the postsynaptic glutamate receptor antagonists. In conclusion, local hyperemia by synaptically-evoked inhibitory neurons was primarily driven by their postsynaptic activities, possibly through NMDA receptor-dependent calcium signaling that was not wholly dependent on nitric oxide.

Adrenergic Receptor Agonists’ Modulation of Dopaminergic and Non-dopaminergic Neurons in the Ventral Tegmental Area

The ventral tegmental area (VTA) neuronal population consists of dopaminergic (DAergic) and non-DAergic neurons (mainly GABAergic), the activity of which is intertwined with VTA behavioral functions. Both DAergic and GABAergic neurons in the VTA have been shown to express adrenergic receptors (ARs) and respond to AR stimulation. The aim of the present study was to demonstrate the effects of selective AR agonists on DAergic and non-DAergic neuronal activity in the central and lateral parts of the VTA using in vivo electrophysiological recording combined with microiontophoretic drug application in anaesthetized rats. Administration of phenylephrine, a selective α1-AR agonist, while having an inhibitory effect on putative DAergic neurons (11% decrease in firing rate), induced a clear excitatory effect (59% increase in firing rate) on putative non-DAergic neurons. In contrast, application of clonidine, a selective α2-AR agonist, or isoprenaline, a selective β-adrenergic receptor agonist, did not change the firing rate of either DAergic or non-DAergic neurons but influenced the firing pattern of non-DAergic cells only. Our results suggest that noradrenaline modulates activity of VTA neurons in vivo primarily via α1, but also via β- and α2-AR to a lesser extent. Furthermore, we show that α1-AR activation has contrasting effects on putative DAergic and non-DAergic neurons. We hypothesize that the phenylephrine-induced inhibition of putative DAergic neurons results from activation of GABAergic terminals present at the site of drug application. Such a mechanism is further supported by the observed α1-AR-induced excitation of putative GABAergic VTA neurons.

THE EFFICACY AND SAFETY OF THE NEW DRUG FISSARIO IN CLINICAL USAGE FOR THE TOPICAL TREATMENT OF THE ACUTE ANAL FISSURE ASSOCIATED WITH CHRONIC HEMORRHOID DISEASE

AIM. To prove primary efficiency of Fissario in comparison with RelifAdvansfor anal fissure therapy and to confirm previously received safety data. MATERIAL AND METHODS. Multicenter, open, randomized, comparative clinical trial in parallel groups with active control (RelifAdvans) was conducted at 18 clinical centers in Russia, 188 patients participated.. Patients with deep and superficial analfissure in combination with chronic hemorrhoids were eligible for participation in trial. Patients administered investigational drugs 2 times a day for 28 days. Efficacy primary endpoint assessed as part ofpatients withfull healing of analfissure and epithelization on Day 28 from start of therapy. Also pharmacokinetics research was performed.: concentration of Nifedipinein blood plasma was assessed. Safety assessment was performed based on frequency and character of registered adverse events. RESULTS. Efficacy analysis showed statistically significant differences in favor of investigational drug in comparison with comparator on efficacy primary endpoint -part of patients with healing of anal fissure at Day 28. Difference of parts of defendants between group of investigational drug and comparator in full data population (with replacement of missed data) was 24,5 % (two-sided confidence interval 95 % for a difference of parts [11,9; 37,0 %], р <0,001), ratio of parts 1,4. Pharmacokinetics analysis based on Nifedipine concentration in plasma showed that observed concentration of Nifedipine in plasma after single rectal and topical use are significantly lower than therapeutic range. Most common adverse events registered during trial were gastrointestinal tract reactions and reactions at investigational drug application site. No serious adverse events, no serious unexpected adverse drug reactions nor cases of death were registered during trial. No influence of Fissario on results of clinical blood tests, biochemical blood tests, general urinalysis and ECG were registered during trial. CONCLUSION. Fissario, ointment for rectal and topical use, is an effective and safe drug for local therapy of acute anal fissure in combination with a chronic hemorrhoids. Fissario achieved more than 40 % superiority in comparison with Relif Advans based on anal fissure epithelizationrate at Day 28 of treatment.

PHYSICAL APPROACH TO TRANSDERMAL DRUG DELIVERY: A REVIEW

Transdermal drug delivery system (TDDS) is the topically administered medication, which when applied to the skin, delivers the drug through the skin for systemic effect at a predetermined and controlled rate. For the drugs, having the short biological half-life, transdermal drug delivery system is convenient. It overcomes the problems like the hepatic first pass effect, improves the compliance of patients, decreases the administration frequency, and reduce the gastrointestinal side effects, peak and valley levels in the blood are avoided. Due to having an advantage of being non-invasive, this delivery has to fulfill some parameters such as high potency, better permeability through skin and non-irritation for better compliance. Skin serves as a site of drug application for local effect as well as systemic effect. There are wide varieties of drugs for which topical or transdermal is a viable option, skin penetration enhancement techniques have been developed for such drugs. There are a number of physical methods to increase drug delivery via the skin, many of which require the use of devices and some of which hold the promise to significantly expand the spectrum of drugs that can be administered transdermally to include water-soluble molecules and macromolecules. Iontophoresis and electroporation represent electrically assisted, physical approaches to enhance delivery of drugs/macromolecules across the stratum corneum. This review highlights the detailed role of physical penetration and describes their mechanism of action. Keywords: transdermal drug delivery system, stratum corneum, skin penetration enhancer, physical enhancer, iontophoresis.

Local treatment of chloasma in pregnant women

Goal. To assess the efficacy and safety of Azelic (15% gel of azelaic acid for topical administration) for the treatment of chloasma in pregnant women. Materials and methods. The study involved 28 pregnant women aged 18-36 (mean age: 24.7) with a normal course of pregnancy. The patients consulted a doctor in the spring, summer or fall complaining of focal skin hyperpigmentation on the face, chin and chest area. Ten patients (35.7%) developed hyperpigmentation prior to their pregnancy and 18 women (64.3%) - during the pregnancy. As of the consultation date, the pregnancy terms in all of the patients were 18-20 weeks. Chloasma was diagnosed by using dermatoscopy and skin examination with the Wood’s lamp. The patients were informed about the content and procedure of the study and gave their consent to take part in the study. Thin layers of Azelic (15% gel of azelaic acid for topical administration) were applied to the hyperpigmented skin of the patients and gently rubbed twice a day (in the morning and evening) as topical treatment for four months. The treatment results were assessed taking into consideration the patient’s subjective assessment, study group structure depending on the clinical efficacy, percentage of adverse events, and survey results based on the Dermatology Life Quality Index questionnaire. To reveal any potential general toxicological effects of the treatment, hepatic samples, total blood count and coagulogram results were analyzed as a part of obstetrical and gynecologic care for pregnant women. Key findings. Positive dynamics of the following characteristics was revealed: subjective assessment of treatment results by the patients, clinical efficacy of treatment and life quality index. Therapeutic results were observed as early as after one month but not later than three months after the treatment began. The therapeutic efficacy was recorded in 92.9-96.4% of all cases after four months of treatment: pigment spots disappeared or became paler, and their dimensions and their contrast as compared to the healthy skin reduced, the skin texture and life quality improved (the Dermatology Life Quality Index reduced from 15.5 ± 1.8 to 6.7 ± 0.5 points against the background of the treatment efficacy: from severe to moderate impact of the disease on the patient’s life, respectively). Five women (17.9%) developed erythema and burning sensations in the drug application site within the first two weeks immediately after the gel application (one of the patients had the same symptoms for six weeks of the treatment); the symptoms were weak and transient and did not require any additional treatment. Hepatic samples, total blood count and coagulogram results were within physiological standards during the treatment. Conclusion. Topical application of Azelic (15% gel of azelaic acid for topical administration) twice a day for four months is an efficient and safe method to treat chloasma in pregnant women.

Percutaneous absorption of salicylates from some commercially available topical products containing methyl salicylate or salicylate salts in rats.

Studies to determine the extent of local tissue penetration of topically applied, commercially available salicylate esters and salts were conducted in male Wistar rats. The salicylate concentration in plasma, tissues underlying the site of drug application, and similar tissues on the contralateral (control) side were measured. The plasma and tissue salicylate levels suggest that direct penetration of salicylate was predominant to the top muscle level on the treated site. Results also suggest that the drugs were first absorbed into the bloodstream and subsequently distributed to both the deeper tissues on the treated site and the contralateral tissues. The topical application of formulations of ester methyl salicylate and salts triethanolamine salicylate and diethylamine salicylate containing comparable salicylate concentrations yielded similar salicylate concentrations in the various tissues. The salicylate concentrations in the deeper tissues approached concentrations observed in the contralateral tissues suggesting that salicylate present in these tissues was due to the systemic blood supply.

Physical and chemical penetration enhancers in transdermal drug delivery system

There is considerable interest in the skin as a site of drug application for both local and systemic effect. However, the skin, in particular the stratum corneum, possesses a formidable barrier to drug penetration thereby limiting topical and transdermal bioavailability. Skin penetration enhancement techniques have been developed to improve bioavailability and to increase the range of drugs for which topical and transdermal delivery is a viable option. The permeation of drug through skin can be enhanced by both chemical penetration enhancement and physical methods. In this review, we have discussed the physical and chemical penetration enhancement technology for transdermal drug delivery as well as the probable mechanisms of action.

Long‐range effects of GABAergic inhibition in gerbil primary auditory cortex

Throughout the literature, the effects of iontophoretically applied neurotransmitter agonists or antagonists on the local activity of neurons are typically studied at the site of drug application. Recently, we have demonstrated long-range inhibitory interactions within the primary auditory cortex (AI) that are effective in complex acoustic situations. To further characterize this long-range functional connectivity, we here report the effects of the inhibitory neurotransmitter gamma-aminobutyric acid (GABA) and the GABA(A) antagonist gabazine (SR 95531) on neuronal activity as a function of distance from the application site reaching beyond the diffusion radius of the applied drug. Neuronal responses to pure tone stimulation were simultaneously recorded at the application site and four additional sites, at distances between 300 and 1350 microm from the application site. We found that whereas application of GABA during best frequency (BF) stimulation in general led to a decrease, and gabazine to an increase, in neuronal activity at the application site, a considerable number of units at remote recording sites showed effects opposite to these local, drug-induced effects. These effects were seen both in spiking activity and in amplitudes of local field potentials. At all locations, the effects varied as a function of pure tone stimulation frequency, pointing to a Mexican-hat-like input function resulting from thalamic inputs to the BF region of the cortical neurons and intracortical interconnections projecting to off-BF regions of the neurons. These data demonstrate the existence of long-range, inhibitory interactions within the gerbil AI, realized either by long-range inhibitory projections or by long-range excitatory projections to local inhibitory interneurons.

Chemical Penetration Enhancers for Transdermal Drug Delivery Systems

Skin as an important site of drug application for both local and systemic effects. However in skin, the stratum corneum is the main barrier for drug penetration. Penetration enhancement technology is a challenging development that would increase the number of drugs availablefor transdermal administration. The permeation of drug through skin can be enhanced by both chemical penetration enhancement and physical methods. In this review, we have discussed the chemical penetration enhancement technology for transdermal drug delivery as well as the probable mechanisms of action.

Synergistically enhanced transdermal permeation and topical analgesia of tetracaine gel containing menthol and ethanol in experimental and clinical studies

The aim of this study is to observe the synergistically enhanced percutaneous penetration and skin analgesia of tetracaine gel containing menthol and ethanol through experimental and clinical studies. Four anesthetic gels containing 4% tetracaine in carbomer vehicle named T-gel (containing no menthol or ethanol), 5%M/T-gel (containing 5% menthol), 70%E/T-gel (containing 70% ethanol, an optimal concentration for antiseptic), and 5%M+70%E/T-gel (containing both 5% menthol and 70% ethanol), respectively, were fabricated. The in vitro mouse skin permeation was investigated using a Franz diffusion cell. The mouse skin morphology was examined by a scanning electron microscope. The in vivo skin analgesic effect in mice was evaluated using the von Frey tests. To determine the efficacy of tetracaine gels for managing the pain in human volunteers, a paralleled, double-blinded, placebo-controlled, randomized controlled trial design combined with verbal pain scores (VPS) was performed. The combination of menthol and ethanol (5%M+70%E/T-gel) conferred significantly higher tetracaine diffusion across full-thickness mouse skin than 5%M/T-gel, 70%E/T-gel, and T-gel. The ultra structure changes of mouse skin stratum corneum treated with 5%M+70%E/T-gel were more marked compared with those of any other tetracaine gel. von Frey tests in mice showed a synergistically enhanced effect of menthol and ethanol on the analgesia of tetracaine gel. The mean VPS were significantly lower for volunteers treated with 5%M+70%E/T-gel than those receiving other gels or the EMLA cream. 5%M+70%E/T-gel possessed the shortest anesthesia onset time, the longest anesthesia duration and the strongest anesthesia efficacy. Seventy percent ethanol in 5%M+70%E/T-gel not only improved the analgesic efficacy of the tetracaine gel through synergistically enhanced percutaneous permeation with menthol but also served as an antiseptic agent keeping drug application site from infection. 5%M+70%E/T-gel is a potential topical anesthesia preparation for clinical use.

Transdermal Drug Delivery: Penetration Enhancement Techniques

There is considerable interest in the skin as a site of drug application both for local and systemic effect. However, the skin, in particular the stratum corneum, poses a formidable barrier to drug penetration thereby limiting topical and transdermal bioavailability. Skin penetration enhancement techniques have been developed to improve bioavailability and increase the range of drugs for which topical and transdermal delivery is a viable option. This review describes enhancement techniques based on drug/vehicle optimisation such as drug selection, prodrugs and ion-pairs, supersaturated drug solutions, eutectic systems, complexation, liposomes, vesicles and particles. Enhancement via modification of the stratum corneum by hydration, chemical enhancers acting on the structure of the stratum corneum lipids and keratin, partitioning and solubility effects are also discussed. The mechanism of action of penetration enhancers and retarders and their potential for clinical application is described.

Preventing the Development of Complex Regional Pain Syndrome after Surgery

COMPLEX regional pain syndrome (CRPS), previously known as reflex sympathetic dystrophy (RSD), is used to describe a syndrome of pain and sudomotor or vasomotor instability. This pain syndrome usually has an initiating noxious event in the periphery, is not limited to the distribution of a single nerve, and is disproportionate to the inciting event. The Consensus Conference of the International Association for the Study of Pain has subclassified CRPS into two forms: CRPS I (formerly RSD) and CRPS II (formerly causalgia). According to the International Association for the Study of Pain, the diagnosis of CRPS I requires (1) continuing pain, allodynia, or hyperalgesia disproportionate to the injury; (2) evidence at some time of edema, changes in skin blood flow, or abnormal sudomotor activity in the region of pain; and (3) no other conditions that would otherwise account for the degree of pain and dysfunction. Motor disturbances and trophic changes, such as altered nail and hair growth, may be observed in some cases. CRPS II is a pain syndrome that starts after a nerve injury and is not necessarily limited to the distribution of the injured nerve. The diagnostic criteria are the same as those of CRPS I. Patients with CRPS I or CRPS II can have sympathetically maintained pain or sympathetically independent pain. Sympathetically maintained pain, a term introduced in 1986 by Roberts, is pathologic pain that is supported by sympathetic efferent activity, circulating catecholamines, and/or increased sensitivity of -adrenergic receptors. Sympathetically maintained pain is identified by the ability to lessen the pain by sympatholytic blocks or interventions. Sympathetically independent pain has components of pain from sources other than sympathetic innervation and is believed to be most commonly observed in advanced cases of CRPS that do not respond to sympathetic blocks. Patients with CRPS may present with components of only sympathetically maintained pain or sympathetically independent pain or, more commonly, a combination of pain from each. Despite increasing research interest, little is known regarding which patients are at increased risk for development of postoperative CRPS and what the optimal perioperative treatment strategy is for those patients undergoing surgery who have a previous history of CRPS. This review outlines the surgical procedures that are believed to increase risk for development of CRPS and describes pharmacologic and regional analgesic techniques that may be of benefit for preventing the development of CRPS after surgery.

Effects of changing the direction of gravity on sector pupil dilations.

To study anterior segment aqueous humor kinetics by using sector pupil dilations with different directions of gravity to change the direction of convectional flow in the anterior chamber. Sector pupil dilations were elicited using focal applications of phenylephrine at the limbus. To vary the direction of gravity (and hence the direction of convectional flow) relative to the eye, subjects' heads were tilted left side down or right side down with gaze horizontal. Pupil elongation and pupil center displacement were measured as functions of time. Early stages of pupil dilation were dominated by pupil elongation toward the site of drug application. Amplitude of elongation was about the same for all the application points, but center displacement depended on the application point. Later stages of pupil dilation showed an interaction between the site of drug application and the direction of gravity. The results are interpreted in terms of anterior chamber convectional flow and its interaction with transpupillary aqueous flow and diffusion. Early distribution in the anterior segment of a substance applied to the limbus appears to be determined primarily by local diffusion, whereas later distribution is affected by movement of the drug in convectional and transpupillary flow.

Activation of barium-sensitive inward rectifier potassium channels mediates remote dilation of coronary arterioles.

Conducted vasodilation seems to be critical for the functional distribution of blood flow in the skeletal muscle microcirculation. However, this vasoregulatory phenomenon has not been documented in the coronary microcirculation, and its underlying mechanism remains elusive. Because potassium ions are potent metabolic vasodilators in the heart, by activating vascular inward rectifier K(+) (K(ir)) channels, we tested the hypothesis that coronary arterioles exhibit remote vasodilation through activation of this type of channel. Porcine coronary arterioles were isolated, cannulated, and pressurized for in vitro study. Vessels dilated concentration-dependently to extraluminal KCl (5 to 20 mmol/L), bradykinin, adenosine, pinacidil, and sodium nitroprusside. A K(ir) channel blocker, BaCl(2) (30 micromol/L), inhibited vasodilatory responses to KCl and bradykinin but not to adenosine, pinacidil, or nitroprusside. In a flow chamber, localized administration of bradykinin, adenosine, and KCl to the downstream end of the arterioles caused approximately 80% dilation at the site of drug application (local site) and also produced 30% to 60% dilation at the upstream end of arterioles (remote site). Nitroprusside produced a similar dilation at the local site but failed to initiate remote vasodilation. In the presence of Ba(2+), adenosine still dilated the local site, but the local dilations to bradykinin and KCl and the remote dilations to adenosine, bradykinin, and KCl were inhibited. We demonstrated that some modes of local vasodilation can be conducted to remote sites in coronary arterioles and that local and remote dilations can occur through different vasodilatory mechanisms. Activation of K(ir) channels seems critical for some agonist-induced local vasodilations and also for the initiation and/or transmission of signals causing remote vasodilation.

Pharmacokinetic and local tissue disposition of [14C]sodium diclofenac following iontophoresis and systemic administration in rabbits

The systemic pharmacokinetics and local drug distribution of sodium diclofenac in skin and underlying tissues was studied. Iontophoresis facilitated local and systemic delivery of diclofenac sodium compared with passive diffusion. The maximum plasma concentration of sodium diclofenac was achieved within 1hof iontophoresis, and the delivery was proportional to applied current density (371±141 and 132±62 μg/L at 0.5 and 0.2 mA/cm2, respectively). The in vivo delivery efficiency for diclofenac in rabbit was 0.15mg/mA·h. The concentrations of sodium diclofenac in the skin, subcutanoeus tissue, and muscle beneath the drug application site (cathode) were significantly greater than plasma concentrations and concentrations of drug in similar tissues at the untreated sites. The results thus suggest that the cutaneous microvasculature is not always a perfect “sink” and that transdermal iontophoresis facilitated the direct penetration of diclofenac sodium to deeper tissues. No skin irritation was observed up to 0.5 mA/cm2 current density and 7mg/mL sodium diclofenac concentration. © 2001 Wiley‐Liss, Inc. and the American Pharmaceutical Association J Pharm Sci 90:1269–1276, 2001

Long-term safety and efficacy of tacrolimus ointment for the treatment of atopic dermatitis in children

Tacrolimus ointment is a nonsteroidal topical immunomodulator that was formulated specifically for the treatment of atopic dermatitis. A total of 255 children, 2 to 15 years of age, with moderate to severe atopic dermatitis applied 0.1% tacrolimus ointment twice daily for up to 12 months to assess long-term safety and efficacy. Patients on average were treated with tacrolimus ointment for 279 days or 87% of study days. Substantial improvements in the signs and symptoms of atopic dermatitis, percent body surface area affected, and the patient's or parent's assessment of pruritus were observed during the first week of treatment and were maintained throughout the study. Transient skin burning and itching were the most common drug application site adverse events. Occurrence of these symptoms decreased after the first few days of treatment. There was no increased incidence of infections or other significant adverse events. Effectiveness of tacrolimus was maintained with prolonged daily use. Tacrolimus ointment (0.1%) is safe and effective for long-term treatment of atopic dermatitis in children. (J Am Acad Dermatol 2001;44:S58-64.)

Hair growth-modulation by adrenergic drugs.

Since we have recently shown that the beta 2-adrenoreceptor (beta 2-AR) expression of selected regions of the hair follicle (HF) epithelium as well as the number of adrenergic nerve fibers in murine skin change in a hair cycle-dependent manner, this has raised the possibility that adrenergic nerves may exert "trophic" functions during HF cycling. To further explore this concept, we have investigated the effect of neuro-pharmacological manipulations on hair growth (anagen) induction in quiescent telogen mouse skin in vivo. Here, we demonstrate that subcutaneous injections of the noradrenaline (NA)-depleting agent guanethidine, or of the neurotoxin 6-hydroxydopamine, but not of the beta 2-AR agonist isoproterenol induce a premature onset of anagen in the lower back skin of C57BL/6 mice. On day 20 after the start of treatment, more than 80% of the guanethidine-treated mice and ca. 65% of the 6-hydroxydopamine-treated (6-OHDA) mice exhibited premature skin darkening and hair growth at the site of drug application, whereas less than one-third of all control animals showed macroscopic signs of anagen development. This was confirmed by histology, demonstrating mature anagen VI HFs only at the immediate site of treatment with guanethidine or 6-OHDA as opposed to resting telogen HFs in the neighboring untreated skin area. This observation further supports the concept that sympathetic nerves are intimately involved in hair growth control and invites one to explore the neuro-pharmacological manipulation of piloneural interactions as a novel therapeutic strategy for the management of hair growth disorders.

Effect of 12-O-tetradecanoyl-phorbol ester and incisional wounding on neuropsin mRNA and its protein expression in murine skin.

The expression of neuropsin mRNA in vivo in mouse skin was examined by in situ hybridization and Northern blotting under stimulated conditions. Two kinds of epidermal stimuli, a topical application of a chemical tumor promoter and incisional wounding, were used. A single topical application of 12-O-tetradecanoyl-phorbol 13-acetate induced epidermal hyperplasia and simultaneously induced an extensive increase in neuropsin mRNA in the suprabasal cells. A full-thickness skin incision also induced a profound increase in neuropsin mRNA in the suprabasal cells surrounding the wound but not in actively proliferating basal cells. The increases in neuropsin mRNA occurred rather late and were limited to the site of drug application or around the incision. Interestingly, neuropsin mRNA was not expressed in the epithelial tongue migrating toward the wound during re-epithelialization. Thus, neuropsin might participate in accelerated epidermal differentiation rather than in the proliferation or migration of keratinocytes in the wound.

Dynamics of axonal microtubules regulate the topology of new membrane insertion into the growing neurites.

Nerve growth depends on the delivery of cell body-synthesized material to the growing neuronal processes. The cellular mechanisms that determine the topology of new membrane addition to the axon are not known. Here we describe a technique to visualize the transport and sites of exocytosis of cell body- derived vesicles in growing axons. We found that in Xenopus embryo neurons in culture, cell body-derived vesicles were rapidly transported all the way down to the growth cone region, where they fused with the plasma membrane. Suppression of microtubule (MT) dynamic instability did not interfere with the delivery of new membrane material to the growth cone region; however, the insertion of vesicles into the plasma membrane was dramatically inhibited. Local disassembly of MTs by focal application of nocodazole to the middle axonal segment resulted in the addition of new membrane at the site of drug application. Our results suggest that the local destabilization of axonal MTs is necessary and sufficient for the delivery of membrane material to specific neuronal sites.