Modulation of synaptic vesicle exocytosis in muscle-dependent long-term depression at the amphibian neuromuscular junction.

We have labeled recycling synaptic vesicles at the somatic Bufo marinus neuromuscular junction with the styryl dye FM2-10 and provide direct evidence for refractoriness of exocytosis associated with a muscle activity-dependent form of long-term depression (LTD) at this synapse. FM2-10 dye unloading experiments demonstrated that the rate of vesicle exocytosis from the release ready pool (RRP) of vesicles was more than halved in the LTD (induced by 20 min of low frequency stimulation). Recovery from LTD, observed as a partial recovery of nerve-evoked muscle twitch amplitude, was accompanied by partial recovery of the refractoriness of RRP exocytosis. Unexpectedly, paired pulse plasticity, another routinely used indicator of presynaptic forms of synaptic plasticity, was unchanged in the LTD. We conclude that the LTD induces refractoriness of the neuromuscular vesicle release machinery downstream of presynaptic calcium entry.

Self-reported risk factors for chlamydia: a survey of pharmacy-based emergency contraception consumers.

OBJECTIVES: To investigate the self-reported risk factors for Chlamydia trachomatis in pharmacy-based emergency contraception (EC) consumers, evaluate their pharmacy experience and determine whether they would be willing to accept a chlamydia test from the pharmacy. METHODS: A survey for women to complete after their EC consultation was developed from themes identified in a literature search. Nineteen pharmacies in the Perth metropolitan region and 13 pharmacies in rural, regional and remote Western Australia (WA) participated in this study. KEY FINDINGS: From the 113 surveys completed (n = 75 from Perth metropolitan; n = 38 from rural, regional and remote WA), 85% of respondents were between 16 and 29 years of age and all (100%) of the women had inconsistent barrier contraception. Almost all (94%) of the women had at least two, and nearly half (47%) had at least three out of the four risk factors for chlamydia. Nearly 70% of the women found it very easy/easy to access a pharmacy and felt very comfortable/comfortable discussing EC with the pharmacist. Significantly more women said they would be willing to accept a chlamydia test from a rural, regional and remote WA pharmacy than from a Perth metropolitan pharmacy (P = 0.003). CONCLUSION: Pharmacy-based EC consumers are at high risk of chlamydia and would be willing to accept a chlamydia test from the pharmacy. There is an urgent need to re-orientate health services in Australia so that all EC consumers, including those obtaining EC from pharmacies, have the opportunity to be tested for chlamydia.

A review of the information-gathering process for the provision of medicines for self-medication via community pharmacies in developing countries.

BACKGROUND: Currently, no review has been completed regarding the information-gathering process for the provision of medicines for self-medication in community pharmacies in developing countries. OBJECTIVE: To review the rate of information gathering and the types of information gathered when patients present for self-medication requests. METHODS: Six databases were searched for studies that described the rate of information gathering and/or the types of information gathered in the provision of medicines for self-medication in community pharmacies in developing countries. The types of information reported were classified as: signs and symptoms, patient identity, action taken, medications, medical history, and others. RESULTS: Twenty-two studies met the inclusion criteria. Variations in the study populations, types of scenarios, research methods, and data reporting were observed. The reported rate of information gathering varied from 18% to 97%, depending on the research methods used. Information on signs and symptoms and patient identity was more frequently reported to be gathered compared with information on action taken, medications, and medical history. CONCLUSION: Evidence showed that the information-gathering process for the provision of medicines for self-medication via community pharmacies in developing countries is inconsistent. There is a need to determine the barriers to appropriate information-gathering practice as well as to develop strategies to implement effective information-gathering processes. It is also recommended that international and national pharmacy organizations, including pharmacy academics and pharmacy researchers, develop a consensus on the types of information that should be reported in the original studies. This will facilitate comparison across studies so that areas that need improvement can be identified.

Provision of primary care to patients with chronic cough in the community pharmacy setting.

BACKGROUND: Community pharmacies are at the forefront of primary care providers and have an important role in the referral of patients to a medical practitioner for review when necessary. Chronic cough is a common disorder in the community and requires medical assessment. The proficiency of community pharmacy staff to refer patients with chronic cough is currently unknown. OBJECTIVE: To assess the ability of community pharmacy staff to recognize and medically refer patients with a chronic nonproductive cough. METHODS: Following ethics approval, a simulated patient study of 156 community pharmacies in Perth, Western Australia, was conducted over a 3-month period. Simulated patients presented to the pharmacy requesting treatment for a cough. The simulated patient required a referral based on a designated scenario. Demographic details, assessment questions, and advice provided were recorded by the simulated patient immediately postvisit. A logistic regression analysis was performed, with referral for medical assessment as the dependent variable. RESULTS: Of the 155 community pharmacies included in the analysis, 38% provided appropriate medical referral. Cough suppressants were provided as therapy in 72% of all visits. Predictors of medical referral were assessment of symptom duration, medical history, current medications being taken, frequency of reliever use, and the position of the pharmacy staff member conducting the consultation. A third of community pharmacies provided appropriate primary care by recommending medical referral advice to patients with chronic cough. The majority of pharmacy staff members acquired information from the patient that suggested a need for medical referral, yet did not provide referral advice. CONCLUSIONS: Appropriate medical referral is more likely when adequate assessment is undertaken and when a pharmacist is directly involved in the consultation. This highlights the need for pharmacies to ensure that processes are in place for patients to access the pharmacist.

Intern pharmacists as change agents to improve the practice of nonprescription medication supply: provision of salbutamol to patients with asthma.

BACKGROUND: Earlier work established an evidence practice gap during provision of nonprescription salbutamol (albuterol). Pharmacist interns are hypothesized to be in a position to improve professional practice in the community pharmacy setting. OBJECTIVE: To explore the potential of intern pharmacists to improve the professional practice of community pharmacy staff in the provision of nonprescription salbutamol. METHODS: Intern pharmacists (n = 157) delivered an asthma intervention in 136 pharmacies consisting of an educational activity to pharmacy staff and a health promotion campaign to consumers. Post-intervention, simulated patients presented to 100 intervention and 100 control community pharmacies with a request for salbutamol. The appropriate outcome was medical referral for poor asthma control and correction of poor inhaler technique. Incidence and quantity of patient assessment and counseling provided during the visit were also assessed. Logistic regression was used to determine the predictors of medical referral. RESULTS: A doubling in the rate of medical referral was seen in the intervention group (19% vs 40%; p = 0.001). Assessment of reliever use frequency was the main predictor of medical referral (OR = 22.7; 95% CI 9.06 to 56.9). Correction of poor inhaler technique did not improve; however, a reduction in salbutamol supplied without patient assessment (23% vs 8%; p = 0.009) or counseling (75% vs 48%; p < 0.001) was noted. CONCLUSIONS: A doubling in the rate of medical referral showed a clear improvement in professional practice during the provision of nonprescription salbutamol. The improved patient outcome in the intervention group was due to increased assessment of reliever use frequency. Identification of poor inhaler technique remained near zero in both groups, which suggests that intern pharmacists were able to improve the current practice of community pharmacies yet were unable to establish a new practice behavior. This study provides evidence that intern pharmacists can act as change agents to improve pharmacy practice.

Measuring the assessment and counseling provided with the supply of nonprescription asthma reliever medication: a simulated patient study.

BACKGROUND: Over one quarter of asthma reliever medications are provided without prescription by community pharmacies in Australia. Evidence that community pharmacies provide these medications with sufficient patient assessment and medication counseling to ensure compliance with the government's Quality Use of Medicines principles is currently lacking. OBJECTIVE: To assess current practice when asthma reliever medication is provided in the community pharmacy setting and to identify factors that correlate with assessment of asthma control. METHODS: Researchers posing as patients visited a sample of Perth metropolitan community pharmacies in May 2007. During the visit, the simulated patient enacted a standardized scenario of someone with moderately controlled asthma who wished to purchase a salbutamol (albuterol) inhaler without prescription. Results of the encounter were recorded immediately after the visit. Regression analysis was performed, with medication use frequency (a marker of asthma control) as the dependent variable. RESULTS: One hundred sixty community pharmacies in the Perth metropolitan area were visited in May 2007. Pharmacists and/or pharmacy assistants provided some form of assessment in 84% of the visits. Counseling was provided to the simulated patients in 24% of the visits. Only 4 pharmacy staff members asked whether the simulated patient knew how to use the inhaler. Significant correlation was found between assessment and/or counseling of reliever use frequency and 3 independent variables: visit length (p < 0.001), number of assessment questions asked (p < 0.001), and the simulated patient who conducted the visit (p < 0.02). CONCLUSIONS: Both patient assessment and medication counseling were suboptimal compared with recommended practice when nonprescription asthma reliever medication was supplied in the community pharmacy setting. Pharmacy and pharmacist demographic variables do not appear to affect assessment of asthma control. This research indicates the need for substantial improvements in practice in order to provide reliever medication in line with Quality Use of Medication principles of ensuring safe and effective use of medication.

Role for the skeletal muscle action potential in non-Hebbian long-term depression at the amphibian (Bufo marinus) neuromuscular junction.

Retrograde signaling from skeletal muscle cells to motor nerve terminals is a recognized mechanism for modulating the strength of neuromuscular transmission. We recently described a form of long-term depression of transmitter release at the mature neuromuscular junction that is dependent on the production of nitric oxide, most likely by the muscle cell (Etherington and Everett 2004 J Physiol (Lond) 559:507-517). We now show that the depression is blocked by treating neuromuscular preparations with mu-conotoxin G111A, an antagonist of skeletal muscle voltage gated sodium channels, indicating that the depression requires postsynaptic action potential firing. Experiments on dually-innervated sartorius muscles revealed that propagation of action potentials generated by low-frequency stimulation of one nerve branch gives rise to nitric-oxide mediated depression at unstimulated nerve terminals located many millimetres away on the same muscle fiber. The non-Hebbian pattern of expression of the depression, as well as its reliance on postsynaptic action potential firing, distinguish it from forms of synaptic depression described at immature neuromuscular synapses and may provide a mechanism for coregulation of the strength of motoneurons innervating the same postsynaptic cell.

Cellular localization of voltage-gated calcium channels and synaptic vesicle-associated proteins in the guinea pig cochlea.

The cellular localization of voltage-gated calcium channels (VGCCs) and synaptic vesicle-associated proteins, SV2, synapsin I, and vesicle-associated membrane protein (VAMP) (synaptobrevin), was investigated in the guinea pig cochlea using immunocytochemistry and confocal laser scanning microscopy. Reactivity, in guinea pig, of antibodies to the alpha1 subunits of L-type, alpha1C [Cav1.2] and alpha 1D [Cav1.3]; P/Q-type, alpha1A [Cav2.1]; and R-type, a1E [Cav2.3] high voltage-activated calcium channels, was determined by Western blotting and immunolabeling of cerebellum. In the cochlea the sensory inner hair cells of the organ of Corti displayed strong intracellular staining, predominantly localized to their basolateral poles, with an antibody directed against the alpha1C subunit. Some alpha1C labeling was also observed in the inner pillar cells, in cell bodies of afferent neurons in the spiral ganglion, and in the inferior region of the spiral ligament. The supporting pillar cells were strongly immunoreactive throughout for alpha1D, but no alpha1D labeling of the inner hair cells was seen. The alpha1A subunit showed a cytoplasmic distribution in all three rows of outer hair cells. alpha1E labeling localized to the outer hair cells, predominantly in the subcuticular plate region, and also to nerve fiber bundles beneath these hair cells. Strong immunoreactivity was consistently seen with antibodies directed against SV2 and synapsin I in neuronal structures surrounding the basolateral surfaces of both the inner and outer hair cells but was absent from the sensory cells themselves. VAMP labeling was found throughout the cytoplasm of the inner hair cells and in neuronal structures beneath the hair cells. These results reveal a differential distribution of VGCC-types in the sensory and nonsensory elements of the guinea pig cochlea, with the inner hair cells expressing alpha1C L-type channels and VAMP but not synapsin I or SV2.

Postsynaptic production of nitric oxide implicated in long-term depression at the mature amphibian (Bufo marinus) neuromuscular junction.

We report here evidence for endogenous NO signalling in long-term (>1 h) synaptic depression at the neuromuscular junction induced by 20 min of 1 Hz nerve stimulation. Synaptic depression was characterized by a 46% reduction in the end-plate potential (EPP) amplitude and a 21% decrease in miniature EPP (MEPP) frequency, but no change to MEPP amplitude, indicating a reduction in evoked quantal release. Both the membrane-impermeant NO scavenger cPTIO and the NOS inhibitor L-NAME blocked depression, suggesting that it is induced by NO originating from a source outside the terminal. The depression was dependent on activation of muscle-type, but not neuronal-type, nAChRs and was still observed when Ca2+ release from the sarcoplasmic reticulum and muscle contraction were blocked with dantrolene. These data suggest that the depression depends on transmission, but not muscle contraction. The calcineurin inhibitors cyclosporin A and FK506, as well as ODQ, an inhibitor of NO-sensitive soluble guanylyl cyclase, Rp-8-pCPT-cGMPS, an inhibitor of cGMP-dependent protein kinase, and the calmodulin antagonist phenoxybenzamine also blocked depression. We propose that low frequency synaptic transmission leads to production of NO at the synapse and depression of transmitter release via a cGMP-dependent mechanism. The NO could be generated either directly from the muscle, or possibly from the Schwann cell in response to an unidentified muscle-derived messenger. We showed that the long-lasting depression of transmitter release was due to sustained activity of the NO signalling pathway, and suggest dephosphorylation of NOS by calcineurin as the basis for continued NO production.

Development of neural tissue and airway smooth muscle in fetal mouse lung explants: a role for glial-derived neurotrophic factor in lung innervation.

We have characterized the distribution of neural tissue and its primary target tissue, airway smooth muscle (ASM), in an in vitro mouse model of early lung development comprising left lung lobes at embryonic Day 12, cultured for 2 or 5 d. Neural tissue was detected with antibodies to protein gene product 9.5 (PGP 9.5), synapsin, and p75NTR (the low-affinity neurotrophin receptor), and smooth muscle with an antibody to alpha-actin. Imaging by confocal microscopy revealed few PGP 9.5-positive neurons at the start of culture; after 2 d clusters of neurons and nerve fibers had appeared along the lobar bronchus and after 5 d along the secondary and tertiary branches. Neural tissue did not just follow the smooth muscle-covered tubules, as seen in vivo, but also grew outside the lobes onto a wide layer of alpha-actin-positive cells, suggesting that smooth muscle may express a trophic factor that attracts nerves. Explants cultured with glial-derived neurotrophic factor (GDNF) exhibited a striking increase in the amount of p75NTR- and PGP 9.5-positive tissue outside the lobes, whereas GDNF-impregnated beads attracted neuronal precursors and influenced the direction of neurite extension. We show that the mouse lung explant is suitable for investigating trophic signals involved in pulmonary innervation and that GDNF may have a role in the early innervation of the developing airways.