Concentrations Of Pilocarpine Research Articles

Transdermal pilocarpine on the skin over salivary glands to increase salivation: an in vivo study

BackgroundHyposalivation is treated using oral cholinergic drugs; however, systemic side effects occasionally lead to discontinuation of treatment. We aimed to investigate the effects of transdermal pilocarpine on the salivary gland skin on saliva secretion and safety in rats.MethodsPilocarpine was administered to rats orally (0.5 mg/kg) or topically on the salivary gland skin (5 mg/body). Saliva volume, the number of sweat dots, and fecal weight were measured along with pilocarpine concentration in plasma and submandibular gland tissues.ResultsSaliva volume significantly increased 0.5 h after oral administration and 0.5, 3, and 12 h after topical administration. Fecal weight and sweat dots increased significantly 1 h after oral administration; however, no changes were observed after topical application. The pilocarpine concentration in the submandibular gland tissues of the topical group was higher than that in the oral group at 0.5, 3, and 12 h of administration.ConclusionsPilocarpine application to salivary gland skin persistently increased salivary volume in rats without inducing sweating or diarrhea. Transdermal pilocarpine applied to the skin over the salivary glands may be an effective and safe treatment option for hyposalivation.

Physiologically Based Pharmacokinetic Modeling and Clinical Extrapolation for Topical Application of Pilocarpine on Eyelids: A Comprehensive Study

Exploiting a convenient and highly bioavailable ocular drug delivery approach is currently one of the hotspots in the pharmaceutical industry. Eyelid topical application is seen to be a valuable strategy in the treatment of chronic ocular diseases. To further elucidate the feasibility of eyelid topical administration as an alternative route for ocular drug delivery, pharmacokinetic and pharmacodynamic studies of pilocarpine were conducted in rabbits. Besides, a novel physiologically based pharmacokinetic (PBPK) model describing eyelid transdermal absorption and ocular disposition was developed in rabbits. The PBPK model of rabbits was extrapolated to human by integrating the drug-specific permeability parameters and human physiological parameters to predict ocular pharmacokinetic in human. After eyelid topical application of pilocarpine, the concentration of pilocarpine in iris peaked at 2 h with the value of 18,724 ng/g and the concentration in aqueous humor peaked at 1 h with the value of 1,363 ng/mL. Significant miotic effect were observed from 0.5 h to 4.5 h after eyelid topical application of pilocarpine in rabbits, while that were observed from 0.5 h to 3.5 h after eyedrop instillation. The proposed eyelid PBPK model was capable of reasonably predicting ocular exposure of pilocarpine after application on the eyelid skin and based on the PBPK model, the human ocular concentration was predicted to be 10-fold lower than that in rabbits. And it was suggested that drugs applied on the eyelid skin could transfer into the eyeball through corneal pathway and scleral pathway. This work could provide pharmacokinetic and pharmacodynamic data for the development of eyelid drug delivery, as well as the reference for clinical applications.

Transdermal iontophoretic application of l-NAME is available in sweating research induced by heat stress in young healthy adults

Iontophoretic transdermal administration of NG-nitro-l-arginine methyl ester hydrochloride [l-NAME, a nitric oxide synthase (NOS) inhibitor] has been used as a non-invasive evaluation of NOS-dependent mechanisms in human skin. However, the availability has yet to be investigated in sweating research. Prior observations using invasive techniques (e.g., intradermal microdialysis technique) to administer l-NAME have implicated that NOS reduces sweating induced by heat stress but rarely influences the response induced by the administration of cholinergic muscarinic receptor agonists. Therefore, we investigated whether the transdermal iontophoretic administration of l-NAME modulates sweating similar to those prior observations. Twenty young healthy adults (10 males, 10 females) participated in two experimental protocols on separate days. Before each protocol, saline (control) and 1% l-NAME were bilaterally administered to the forearm skin via transdermal iontophoresis. In protocol 1, 0.001% and 1% pilocarpine were iontophoretically administered at l-NAME-treated and untreated sites. In protocol 2, passive heating was applied by immersing the lower limbs in hot water (43 °C) until the rectal temperature increased by 0.8 °C above baseline. The sweat rate was continuously measured throughout both protocols. Pilocarpine-induced sweat rate was not significantly different between the control and l-NAME-treated sites in both pilocarpine concentrations (P ≥ 0.316 for the treatment effect and interaction of treatment and pilocarpine concentration). The sweat rate during passive heating was attenuated at the l-NAME-treated site relative to the control (treatment effect, P = 0.020). Notably, these observations are consistent with prior sweating studies administrating l-NAME into human skin using intradermal microdialysis techniques. Based on the similarity of our results with already known observations, we conclude that transdermal iontophoresis of l-NAME is a valid non-invasive technique for the investigation of the mechanisms of sweating related to NOS during heat stress.

Potential antiproliferative and apoptotic effects of pilocarpine combined with TNF alpha in chronic myeloid leukemia cells.

Pilocarpine is a selective M1/M3 agonist of muscarinic acetylcholine receptor subtypes. Muscarinic acetylcholine receptors are G protein-coupled receptors. These receptors are different drug targets. The aim of the present work was to investigate the effect of pilocarpine on the expression of M3 muscarinic acetylcholine receptor, the AChE activity, IL-8 release response, and proliferation in K562 cells, via muscarinic receptor activation.Human chronic myeloid leukemic cell cultures were incubated with drugs. Proliferation assays were performed by BrdU assay. Expression of M3 muscarinic acetylcholine receptor and apoptosis proteins such as bcl, bax, cyt C, and caspases was assessed with the semiquantitative Western blotting method.Pilocarpine inhibits chronic myeloid cell proliferation and M3 muscarinic acetylcholine receptor protein expression. Pilocarpine increases caspase-8 and -9 expression levels, upregulating the proapoptotic protein Bax and downregulating the expression levels of the antiapoptotic protein Bcl-2. The apoptotic activity of pilocarpine is associated with an increase in AChE activity. M3 muscarinic acetylcholine receptors can activate multiple signal transduction systems and mediate inhibitory effects on chronic myeloid K562 cell proliferation depending on the presence of 1% FBS conditions. This apoptotic effect of pilocarpine may be due to the concentration of pilocarpine and the increase in AChE level.Our results suggest that inhibition of cell proliferation by inducing apoptosis of pilocarpine in K562 cells may be one of the targets. M3 selective agonist may have therapeutic potential in chronic myeloid leukemia.

Application of Green Technologies in Design-Based Extraction of Celastrus paniculatus (Jyotishmati) Seeds, SEM, GC-MS Analysis, and Evaluation for Memory Enhancing Potential.

BackgroundThe Celastrus paniculatus (CP), commonly known as Jyotishmati, is considered as “elixir of life” by Indian people for the prevention or management of many ailments. The seed powder and its extract have widely used commercially for the preparation of various Ayurvedic formulations for the improvement of memory. CP seeds were generally extracted by conventional extraction methods (CEMs) which are assumed to impact environment burden and also produce low extract yield. Green extraction with possible improvement in extract yield has always been the need of hour for selected medicinal plant.ObjectiveIn the present research, we aimed to optimize the different extraction factors in microwave and ultrasound-based extraction. The various extracts obtained in conventional and green methods are also evaluated for the possible improvement in memory enhancing potential.Materials and MethodsThe selected medicinal herb was extracted by CEM (maceration and percolation). In green methods such microwave-assisted extraction (MAE) and ultrasound assisted-extraction (UAE), various parameters were optimized using Box-Behnken design coupled with response surface methodology. The scanning electron microscopy (SEM) and gas chromatography–mass spectroscopy (GC-MS) analyses were also done to confirm the possible improvement in concentration of plant actives. The Swiss albino mice were used to evaluate memory enhancing potential of different extracts.ResultsAt the optimized conditions MAE and UAE the extraction yield, total phenolic content (TPC) and Total flavonoid content (TFC) are significantly improved. The GC-MS analysis further confirms the improvement in concentration of certain fatty acid esters, pilocarpine, and steroidal compounds in optimized extracts. The optimized extracts also exhibited the significant improvement in behavioral parameters, oxidative stress-induced parameters, and acetylcholinesterase inhibitory potential.Discussion and ConclusionFrom the results, we can say that the application of green technologies in design-based extraction of selected herb not only significantly reduces the extraction time but also improves the extract yield and concentration of plant actives. In nutshell, it can be concluded that the green approaches for extraction of seeds of Celastrus paniculatus could be scale up at a commercial level to meet the rising demand for herbal extract.

Topical Xerostomia Treatment with Hyaluronate Sheets Containing Pilocarpine.

Sjogren's syndrome and radiation therapy for head and neck cancers are often accompanied by xerostomia. Oral pilocarpine (PCP) to treat xerostomia produces systemic side effects, such as runny nose and lacrimation. To improve the therapeutic efficacy of PCP and reduce the aforementioned side effects, we developed a topical delivery system for PCP using freeze-dried sheets of hyaluronic acid (HA). The advantages of HA sheets over conventional oral formulations were examined through in vivo pharmacokinetic and pharmacodynamic studies after their application to oral tissues and salivary glands. The concentration of PCP in the submucosal tissue of the oral cavity was determined using the microdialysis (MD) method after buccal application of HA sheets containing PCP to hamsters. The concentration of PCP in the MD outflow was quite low after gastric administration, whereas the PCP concentration in plasma was high. In contrast, after buccal application of HA sheets containing PCP, the concentration of the drug in the MD outflow increased, despite the negligible concentration in plasma. These findings indicated that both enhancement of saliva secretion and the avoidance of systemic side effects could be achieved through buccal administration of PCP-loaded HA sheets. In addition, the pharmacodynamic study showed that when compared with intravenous and gastric administration, salivary application of HA sheets containing PCP resulted in similar volumes of saliva secretion and reduced lacrimal secretions. In conclusion, freeze-dried HA sheets allow for the development of a novel buccal delivery system with enhanced therapeutic efficacy and safety to treat xerostomia.

N-Methyl-(2S, 4R)-trans-4-hydroxy-L-proline, the major bioactive compound from Sideroxylon obtusifolium, attenuates pilocarpine-induced injury in cultured astrocytes

Glial cells have been implicated in temporal lobe epilepsy in humans and in its models. Astrocytes are lost in several brain regions after acute seizures induced by pilocarpine and may suffer hyperplasia at subsequent time points. This study investigated the effect of N-methyl-(2S,4R)-trans-4-hydroxy-L-proline (NMP) on astrocytes exposed to cytotoxic concentrations of pilocarpine. Astrocytes were incubated with pilocarpine (half maximal inhibitory concentration (IC50)=31.86 mM) for 24 h. Afterwards, they were treated with NMP at concentrations ranging from 3.12 to 100 μg/mL for 24 h. Cell viability was assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cytoplasmic reactive oxygen species (ROS) and mitochondrial transmembrane potential (ΔΨm) were analyzed by flow cytometry using 2',7'-dichlorofluorescein diacetate (DCFH-DA) and rhodamine-123 (Rho123), respectively. Expression of glial fibrillary acidic protein (GFAP) and voltage-dependent anion channel-1 (VDAC-1) were measured by western blot. Pilocarpine significantly decreased cell viability and mitochondrial potential and increased ROS concentration significantly by 6.7 times compared to the control. NMP concentrations ≥25 µg/mL protected astrocytes against pilocarpine-induced injury in a concentration-dependent manner. Concomitantly, NMP reduced cytoplasmic ROS accumulation to 27.3, 24.8, and 12.3% in the groups treated with 25, 50, and 100 µg/mL NMP, respectively. NMP also protected mitochondria from pilocarpine-induced depolarization. These effects were associated with improvement of pilocarpine-induced GFAP and VDAC-1 overexpression, which are important biomarkers of astrocyte dysfunction. In conclusion, the improvement of ROS accumulation, VDAC-1 overexpression, and mitochondrial depolarization are possible mechanisms of the NMP protective action on reactive astrocytes.

Adie's Pupil: A Diagnostic Challenge for the Physician.

Adie’s pupil, also called tonic pupil, is mainly seen in young women. Most patients have unilateral eye involvement. The pupil of the affected side is significantly larger than that on the healthy side. The direct and indirect light reflection from the pupil on the affected side disappears. The pupil on the affected side is sensitive to low concentrations of pilocarpine. The pathogeneses of Adie’s pupil are complex, some of which are insidious and lack corresponding specific diseases. Through a literature review, we found that Adie’s pupil is mainly associated with infectious diseases, most commonly syphilis, followed by immune diseases and paraneoplastic syndromes. The ophthalmological symptoms and pupil abnormalities can disappear after active treatment of the primary disease. Pilocarpine can be used to treat ophthalmologic symptoms, such as blurred vision, for which patients might visit an ophthalmologist or neurologist. It is essential for clinicians to improve their understanding of the disease to avoid misdiagnosis. Differential diagnosis between Adie’s pupil, oculomotor nerve palsy, anticholinergic drug overdose, Argyll-Robertson pupil, and congenital mydriasis need to be identified by the physician. Here, the clinical manifestations, pathogenesis, relationship between Adie’s pupil and diseases, and differential diagnosis of Adie’s pupil are reviewed.

Dilute pilocarpine test for diagnosis of Adie\u2019s tonic pupil

We have compared the diagnostic ability of different concentrations of 0.125% and 0.0625% dilute pilocarpine for detecting denervation supersensitivity in unilateral Adie’s tonic pupil. This retrospective, observational, case–control study involved 117 subjects, consisting of 56 patients with unilateral Adie’s tonic pupil and 61 controls with other causes of unilateral dilated pupils. Subjects underwent the dilute pilocarpine test with one of the two concentrations, 0.125% or 0.0625%. Pupillary light reflex was recorded with a dynamic pupillometer at baseline and at 30–40 min after instilling one of the two concentrations of dilute pilocarpine. Diagnostic accuracy of two different concentrations of the dilute pilocarpine test, 0.125% group versus 0.0625% group, were compared by area under the receiver operating characteristic curve (AUC). Diagnostic ability of the dilute pilocarpine test for detecting denervation supersensitivity in unilateral Adie’s tonic pupil was significantly better in the 0.0625% group than in the 0.125% group (AUC = 0.954 vs. 0.840, respectively, P = 0.047). In the 0.0625% group, the change in maximal pupil diameter of ≥ 0.5 mm after topical pilocarpine instillation showed 100% sensitivity and 82.8% specificity for detecting Adie’s tonic pupil. This study confirmed that pupillary constriction with 0.0625% pilocarpine is better than 0.125% pilocarpine for detecting denervation supersensitivity in Adie’s tonic pupil. Digital pupillometry is a reliable method for assessing denervation supersensitivity in Adie's tonic pupil.

Effects of L-type voltage-gated Ca2+ channel blockade on cholinergic and thermal sweating in habitually trained and untrained men.

We evaluated the hypothesis that the activation of L-type voltage-gated Ca2+ channels contributes to exercise training-induced augmentation in cholinergic sweating. On separate days, 10 habitually trained and 10 untrained men participated in two experimental protocols. Prior to each protocol, we administered 1% verapamil (Verapamil, L-type voltage-gated Ca2+ channel blocker) and saline (Control) at forearm skin sites on both arms via transdermal iontophoresis. In protocol 1, we administered low (0.001%) and high (1%) doses of pilocarpine at both the verapamil-treated and verapamil-untreated forearm sites. In protocol 2, participants were passively heated by immersing their limbs in hot water (43°C) until rectal temperature increased by 1.0°C above baseline resting levels. Sweat rate at all forearm sites was continuously measured throughout both protocols. Pilocarpine-induced sweating in Control was higher in trained than in untrained men for both the concentrations of pilocarpine (both P ≤ 0.001). Pilocarpine-induced sweating at the low-dose site was attenuated at the Verapamil versus the Control site in both the groups (both P ≤ 0.004), albeit the reduction was greater in trained as compared with in untrained men (P = 0.005). The verapamil-mediated reduction in sweating remained intact at the high-dose pilocarpine site in the untrained men (P = 0.004) but not the trained men (P = 0.180). Sweating did not differ between Control and Verapamil sites with increases in rectal temperature in both groups (interaction, P = 0.571). We show that activation of L-type voltage-gated Ca2+ channels modulates sweat production in habitually trained men induced by a low dose of pilocarpine. However, no effect on sweating was observed during passive heating in either group.

Multifunctional glutathione-dependent hydrogel eye drops with enhanced drug bioavailability for glaucoma therapy

The development of drug carriers with advanced functions to overcome the low bioavailability of conventional eye drops is critical for the effective treatment of chronic ocular diseases. This paper reports a functional strategy for developing a new hydrogel eye drop with strong mucoadhesive, tight-junction opening, and antioxidant properties, which play important roles in improving the topical treatment of glaucoma. The eye-drop carrier was rationally designed through the conjugation of gelatin with poly(N-isopropylacrylamide) (GN) via the formation of amide linkages, followed by free-radical grafting with glutathione (GSH). In vitro studies revealed that the GN-GSH hydrogels had good ocular biocompatibility, high antioxidant activity, and high paracellular permeability. Interestingly, grafting an appropriate amount of GSH to the GN backbone led to a tradeoff between the bioadhesion and biodegradation properties of the hydrogel, which prolonged its retention on the conjunctival sac of a rabbit’s eye without causing any ocular discomfort/irritation. In a rabbit glaucoma model, pharmacological treatment with a single-dose topical instillation of the optimized pilocarpine-loaded GN-GSH hydrogel effectively suppressed disease progression for 14 d, whereas that employing only a GN-based formulation mitigated glaucoma development for 3 d (modest efficacy). This significant enhancement is ascribed to the improved drug bioavailability achieved via the rational exploitation of GSH, which resulted in an eightfold increase in the pilocarpine concentration in the aqueous humor of the glaucomatous eyes. These findings can be valuable for the development of multifunctional GSH-dependent hydrogel eye drops as long-acting ophthalmic formulations for the efficient management of complex and chronic ocular diseases.

Pharmacokinetics and Tissue Distribution of Pilocarpine After Application to Eyelid Skin of Rats

Extending the delivery of drugs into the eyes while reducing systemic bioavailability is of utmost importance in the management of chronic ocular diseases. Topical application onto the lower eyelid skin, as an alternative to eye drops, is seen to be a valuable strategy in the treatment of chronic eye diseases. To elucidate the critical value of delivering drugs in solution onto the eyeball through the eyelid skin, pharmacokinetic studies of pilocarpine were conducted, and the results were verified using a direct pharmacodynamic study in rats. The mean residence time of pilocarpine after topical eyelid application to the eyelid skin, conjunctiva, eyeball, and plasma were 14.9, 8.50, 6.29, and 8.11 h, respectively. Conjunctiva and eyeball concentrations of pilocarpine at 8 h were 80-fold and 8-fold higher after topical eyelid application, respectively, than those for eye drops. Pupillary constriction was sustained over 8 h after topical eyelid application. Topical eyelid skin application exhibited a localized drug absorption and specific drug accumulation in the ocular tissues. Hence, it is rational to prepare topical formulations directed onto the eyelid skin, which is suitable for drugs required for long-term treatment.

New low-dose liquid pilocarpine formulation for treating dry mouth in Sj\xf6gren\u2019s syndrome: clinical efficacy, symptom relief, and improvement in quality of life

BackgroundPatients with Sjögren’s syndrome (SS) typically present clinically with xerostomia (dry mouth) because of progressive damage to the exocrine glands. We developed a new, low-dose pilocarpine/sodium alginate (LPA) solution with pilocarpine hydrochloride to inhibit systemic adverse effects by administering via the oral mucosa. The purpose of this study was to assess its stability, safety, and efficacy.MethodsThe pilocarpine concentration in an LPA liquid formulation was measured 3, 7, 14, and 28 days after preparation to assess its stability. A prospective clinical trial was undertaken to assess the efficacy and safety of the LPA solution as a symptomatic treatment for dry mouth in SS. Patients (n = 24) with clinically significant xerostomia were enrolled after providing written informed consent. Whole-mouth salivary flow rate was measured twice; immediately before and 60 min after LPA application. Symptoms were assessed by questionnaire with visual analog scales or checkboxes before the first application (baseline), and then once daily for 7 days.ResultsThe pilocarpine content 3, 7, 14, and 28 days after preparation showed no marked change, confirming its stability. Salivary flow was significantly increased from 0.076 ± 0.092 g/30 s to 0.122 ± 0.140 g/30 s 60 min after LPA administration (P < 0.001). Dry mouth and thirstiness showed significant improvement compared with that of baseline (P ≤ 0.01). The only adverse effect was sweating, and no serious drug-related adverse events were reported.ConclusionsThis new, low-dose pilocarpine formulation was well-tolerated and resulted in significant improvements in symptoms of dry mouth and other xerostomic conditions in patients with SS.Trial registrationThe study approval number in the institution; 08–068-2. Registered January 19, 2009. UMIN000029307. Registered 27 September 2017 (retrospectively registered).

Structural determinants at the M2 muscarinic receptor modulate the RGS4-GIRK response to pilocarpine by impairment of the receptor voltage sensitivity

Membrane potential controls the response of the M2 muscarinic receptor to its ligands. Membrane hyperpolarization increases response to the full agonist acetylcholine (ACh) while decreasing response to the partial agonist pilocarpine. We previously have demonstrated that the regulator of G-protein signaling (RGS) 4 protein discriminates between the voltage-dependent responses of ACh and pilocarpine; however, the underlying mechanism remains unclear. Here we show that RGS4 is involved in the voltage-dependent behavior of the M2 muscarinic receptor-mediated signaling in response to pilocarpine. Additionally we revealed structural determinants on the M2 muscarinic receptor underlying the voltage-dependent response. By electrophysiological recording in Xenopus oocytes expressing M2 muscarinic receptor and G-protein-gated inwardly rectifying K+ channels, we quantified voltage-dependent desensitization of pilocarpine-induced current in the presence or absence of RGS4. Hyperpolarization-induced desensitization of the current required for RGS4, also depended on pilocarpine concentration. Mutations of charged residues in the aspartic acid-arginine-tyrosine motif of the M2 muscarinic receptor, but not intracellular loop 3, significantly impaired the voltage-dependence of RGS4 function. Thus, our results demonstrated that voltage-dependence of RGS4 modulation is derived from the M2 muscarinic receptor. These results provide novel insights into how membrane potential impacts G-protein signaling by modulating GPCR communication with downstream effectors.

Influence of Pilocarpine and Timolol on Human Meibomian Gland Epithelial Cells.

Investigators have discovered that topical antiglaucoma drugs may induce meibomian gland dysfunction. This response may contribute to the dry eye disease commonly found in patients with glaucoma taking such medications. We hypothesize that drug action involves a direct effect on human meibomian gland epithelial cells (HMGECs). To test this hypothesis, we examined the influence of the antiglaucoma drugs, pilocarpine and timolol, on the morphology, survival, proliferation, and differentiation of HMGECs. Immortalized (I) HMGECs (n = 2-3 wells/treatment/experiment) were cultured with multiple concentrations of pilocarpine or timolol for up to 7 days. Experiments included positive controls for proliferation (epidermal growth factor and bovine pituitary extract) and differentiation (azithromycin). Cells were enumerated using a hemocytometer and evaluated for morphology, neutral lipid staining, and lysosome accumulation. Our results demonstrate that pilocarpine and timolol cause a dose-dependent decrease in the survival of IHMGECs. The clinically used concentrations are toxic and lead to cell atrophy, poor adherence, or death. By contrast, drug levels that are known to accumulate within the conjunctiva, adjacent to the meibomian glands, do not influence IHMGEC survival. These latter concentrations also have no effect on IHMGEC proliferation or differentiation, and they do not interfere with the ability of azithromycin to stimulate cellular neutral lipid and lysosome accumulation. This dose of pilocarpine, though, did suppress the epidermal growth factor+bovine pituitary extract-induced proliferation of IHMGECs. Our results support our hypothesis and demonstrate that these antiglaucoma drugs, pilocarpine and timolol, have direct effects on HMGECs that may influence their morphology, survival, and proliferative capacity.

On the importance of Bloom number of gelatin to the development of biodegradable in situ gelling copolymers for intracameral drug delivery

To overcome the drawbacks associated with conventional antiglaucoma eye drops, this work demonstrated the feasibility of an effective alternative strategy to administer pilocarpine directly via intracameral injections of drug-containing biodegradable in situ gelling GN copolymers composed of gelatin and poly(N-isopropylacrylamide). Specifically, this study aims to understand the importance of Bloom number of gelatin, a physicochemical parameter, to the development of GN carriers for intracameral drug delivery in glaucoma therapy. Our results showed that both imino acid and triple-helix contents increased with increasing Bloom index from 75⿿100 to 300. The drug encapsulation efficiency in response to temperature-triggered phase transition in GN copolymers was affected by the Bloom index of gelatin. In addition, the differences in protein secondary structure significantly influenced the degradation rates of GN carriers, which were highly correlated with drug release profiles. The increase in released pilocarpine concentration led to a high intracellular calcium level in rabbit ciliary smooth muscle cell cultures, indicating a beneficial pharmacological response to a drug. Irrespective of Bloom number of gelatin, all carrier materials exhibited excellent in vitro and in vivo biocompatibility with corneal endothelium. In a glaucomatous rabbit model, intracameral injections of pilocarpine-containing GN synthesized from gelatins with various Bloom numbers had different abilities to improve ocular hypertension and induce pupillary constriction, indicating distinct antiglaucoma efficacies due to in vivo drug release. It is concluded that the effects on pharmacological treatment using GN carriers for intracameral pilocarpine administration demonstrate a strong dependence on the Bloom number of gelatin.

Flight and Walking in Locusts–Cholinergic Co-Activation, Temporal Coupling and Its Modulation by Biogenic Amines

Walking and flying in locusts are exemplary rhythmical behaviors generated by central pattern generators (CPG) that are tuned in intact animals by phasic sensory inputs. Although these two behaviors are mutually exclusive and controlled by independent CPGs, leg movements during flight can be coupled to the flight rhythm. To investigate potential central coupling between the underlying CPGs, we used the muscarinic agonist pilocarpine and the amines octopamine and tyramine to initiate fictive flight and walking in deafferented locust preparations. Our data illustrate that fictive walking is readily evoked by comparatively lower concentrations of pilocarpine, whereas higher concentrations are required to elicit fictive flight. Interestingly, fictive flight did not suppress fictive walking so that the two patterns were produced simultaneously. Frequently, leg motor units were temporally coupled to the flight rhythm, so that each spike in a step cycle volley occurred synchronously with wing motor units firing at flight rhythm frequency. Similarly, tyramine also induced fictive walking and flight, but mostly without any coupling between the two rhythms. Octopamine in contrast readily evoked fictive flight but generally failed to elicit fictive walking. Despite this, numerous leg motor units were recruited, whereby each was temporarily coupled to the flight rhythm. Our results support the notion that the CPGs for walking and flight are largely independent, but that coupling can be entrained by aminergic modulation. We speculate that octopamine biases the whole motor machinery of a locust to flight whereas tyramine primarily promotes walking.

Cevimeline-Induced Monophasic Salivation from the Mouse Submandibular Gland: Decreased Na+Content in Saliva Results from Specific and Early Activation of Na+/H+Exchange

Cevimeline and pilocarpine are muscarinic agonists used clinically to treat dry mouth. In this study, we explored fluid secretion from mouse submandibular glands to determine the mechanism of cevimeline, pilocarpine, and an experimentally used agent carbachol. Cevimeline evoked almost the same amount of secretion at concentrations from 30 μM to 1 mM. Pilocarpine also induced secretion at a concentration as low as 1 μM and was the most powerful secretagogue at 10 μM. Secretion was induced by carbachol at 0.1 μM, with maximum secretion at 1.0 μM. Cevimeline induced monophasic secretion at all concentrations tested, whereas higher concentrations of pilocarpine and carbachol induced secretion with variable kinetics, i.e., an initial transient high flow rate, followed by decreased secretion after 2 to 3 min. In the presence of an epithelial Na(+) channel blocker, amiloride, neither carbachol nor pilocarpine affected the Na(+) level of secreted saliva; however, it significantly increased the Na(+) content of cevimeline-induced saliva. The intracellular Ca(2+) response of acinar cells was almost identical among all three agents, although recovery after drug removal was slower for cevimeline and pilocarpine. A profound decrease in intracellular pH was observed during pilocarpine and carbachol treatment, whereas intracellular acidification induced by cevimeline was only seen in the presence of a Na(+)/H(+) exchange inhibitor. When external HCO(3)(-) was removed, cevimeline-induced saliva significantly decreased. These findings suggest that cevimeline specifically activates Na(+)/H(+) exchange and may promote Na(+) reabsorption by stabilizing epithelial sodium channel activity.

Aging and topical pilocarpine concentrations effects on pupil size and tear flow rate

Purpose This is to evaluate the effects of age and different concentrations of pilocarpine on tear flow rate and pupil size. Methods The tear flow rates and pupil sizes of eighty-one volunteers were measured with Schimer's strips and a meter rule gauge, respectively. These procedures were also employed in testing the effects of 2 % and 4 % concentrations of pilocarpine on tear secretion and pupil constriction. Results There were significant differences between the initial mean tear flow rate and the mean tear flow rates after the installation of 2 % pilocarpine (critical t = 1.96 and calculated t = 6.46) and 4 % pilocarpine (calculated t = 8.83). There were no significant differences in the initial tear flow rates between the sexes. There were significant differences between the mean tear flow rates obtained with 2 % and 4 % concentrations of pilocarpine (calculated t = 3.41). The mean initial tear flow rate for the eighty-one volunteers was 13.84 mm ± 1.19 (SD).There were significant differences between the mean initial pupil size 2.96 mm ± 1.04 (SD) and the mean pupil sizes 2.38 mm ± 1.11(SD) after the instillation of 2 % pilocarpine (Critical t = 1.96 and calculated t = 2.46). Conclusion The basal tear flow rate decreased with advancing age and topical pilocarpine increased tears secretion in all the age groups. The age groups (31 years and above) were more tolerant to the stinging effect of pilocarpine and induced reduced pupil sizes, but there were no significant differences between the mean pupil sizes obtained with the pilocarpine concentrations.

Topical and intravenous pilocarpine stimulated accommodation in anesthetized rhesus monkeys

Many studies have used pilocarpine to stimulate accommodation in both humans and monkeys. However, the concentrations of pilocarpine used and the methods of administration vary. In this study, three different methods of pilocarpine administration are evaluated for their effectiveness in stimulating accommodation in rhesus monkeys. Experiments were performed in 17 iridectomized, anesthetized rhesus monkeys aged 4–16 years. Maximum accommodation was stimulated in all these monkeys with a 2% pilocarpine solution maintained on the cornea for at least 30 min in a specially designed perfusion lens. In subsequent topical pilocarpine experiments, baseline refraction was measured with a Hartinger coincidence refractometer and then while the monkeys were upright and facing forward, commercially available pilocarpine (2, 4, or 6%) was applied topically to the cornea as 2 or 4 drops in two applications or 6 drops in three applications over a five minute period with the eyelids closed between applications. Alternatively, while supine, 10–12 drops of pilocarpine were maintained on the cornea in a scleral cup for 5 min. Refraction measurements were begun 5 min after the second application of pilocarpine and continued for at least 30 min after initial administration until no further change in refraction occurred. In intravenous experiments, pilocarpine was given either as boluses ranging from 0.1 mg/kg to 2 mg/kg or boluses followed by a constant infusion at rates between 3.06 mg/kg/h and 11.6 mg/kg/h. Constant 2% pilocarpine solution on the eye in the perfusion lens produced 10.88 ± 2.73 D (mean ± SD) of accommodation. Topically applied pilocarpine produced 3.81 D ± 2.41, 5.49 D ± 4.08, and 5.55 D ± 3.27 using 2%, 4%, and 6% solutions respectively. When expressed as a percentage of the accommodative response amplitude obtained in the same monkey with constant 2% pilocarpine solution on the eye, the responses were 34.7% for 2% pilocarpine, 48.4% for 4% pilocarpine, and 44.6% for 6% pilocarpine. Topical 4% and 6% pilocarpine achieved similar, variable accommodative responses, but neither achieved maximum accommodation. IV boluses of pilocarpine achieved near maximal levels of accommodation at least ten times faster than topical methods. Doses effective for producing maximum accommodation ranged from 0.25 mg/kg to 1.0 mg/kg. IV pilocarpine boluses caused an anterior movement of the anterior lens surface, a posterior movement of the posterior lens surface, and a slight net anterior movement of the entire lens. Considerable variability in response amplitude occurred and maximum accommodative amplitude was rarely achieved with topical application of a variety of concentrations of commercially available pilocarpine. Intravenous infusion of pilocarpine was a rapid and reliable method of producing a nearly maximal accommodative response and maintaining accommodation when desired.