• Journal of Pharmaceutical Investigation
• /
• v.24 no.3
• /
• pp.49-57
• /
• 1994

The objective of this study was to determine whether Pz-peptide, an enhancer of hydrophilic solute permeability in the intestine, could elevate the paracellular permeability of the cornea and conjunctiva in the pigmented rabbit. The in vitro penetration of four hydrophilic solutes, mannitol (MW 182), fluorescein (MW 376), FD-4 (FITC-dextran, 4 KDa), and FD-10 (FITC-dextran, 10 KDa) across the pigmented rabbit cornea and conjunctiva was studied either in the presence or absence of 3 mM enhancers. Drug penetration was evaluated using the modified Ussing chamber. The conjunctiva was more permeable than the cornea to all four markers. EDTA and cytochalasin B showed higher effects on marker transport than Pz-peptide, but Pz-peptide elevated the corneal transport of mannitol, fluoresein, and FD-4 by 50%, 26%, and 50%, respectively, without affecting FD-10 transport. Possibly due to the leakier nature of the conjunctiva, 3 mM Pz-peptide elevated the transport of only FD-4 by about 45%, without affecting the transport of other markers. Furthermore, the transport of Pz-peptide itself across the cornea and conjunctiva increased with increasing concentration in the 1-5 mM range, suggesting that Pz-peptide enhanced its own permeability, possibly by elevating paracellular permeability. Effects of ion transport inhibitors on Pz-peptide transport were then investigated. PZ-peptide penetration was not changed by mucosal addition of $10\;{\mu}M$ amiloride or $10\;{\mu}M$ hexamethylene amiloride, inhibiting serosal $Na^{+}$ exit by $100\;{\mu}M$ ouabain, or replacing $Na^{+}$ with choline chloride in the mucosal side buffer. These results seggested that Pz-peptide enhanced the paracellular permeability of rabbit cornea and conjunctiva and further indicate that ion transporters were not involved in the Pz-peptide induced elevation of paracellular marker permeability.

• Journal of Pharmaceutical Investigation
• /
• v.26 no.1
• /
• pp.43-53
• /
• 1996

The objective of this study was to determine whether 4-phenylazobezyloxycarbonyl-Pro-Leu-Gly-Pro-D-Arg (Pz-peptide), an enhancer of hydrophilic solute permeability in the intestine, could elevate the paracellular permeability of hydrophilic drugs across cornea and conjunctiva in the rabbit. The in-vitro penetration of hydrophilic drugs (mannitol, atenolol) and lipophilic drug (propranolol) across the rabbit cornea and conjunctiva was studied either in the presence or absence of 3 mM Pz-peptide. Drug penetration was evaluated using the modified Ussing chamber. The conjunctiva was more permeable than the cornea to all drugs. Pz-peptide showed enhanced effects on the drug transport across cornea and conjunctiva in a concentration dependent manner. Effects or ion transport inhibitor on the mannitol penetration were then investigated. Mannitol penetration was not changed by serosal addition of $100\;{\mu}M$ ouabain, suggesting that $Na^+/K^+$ ion tranporter was not involved in the Pz-peptide induced elevation of paracellular drug permeability. Furthermore, effects of Pz-peptide and EDTA on the transport of atenolol and propranolol into the ocular tissues or blood circulation after its administration into both eyes were investigated. EDTA showed enhanced effect on propranolol transport into the ocular tissues, but Pz-peptide did not show significant difference. Systemic absorption of propranolol by the addition of EDTA or Pz-peptide was not changed. On the other hand, EDTA and Pz-peptide elavated the atenolol transport into the ocular tissues. The transport of atenolol into the blood circulation was also enhanced by the addition of EDTA, but no effect was observed by the addition of Pz-peptide. The above findings suggest that Pz-peptide would be used as an paracellular pathway enahncer of hydrophilic drugs into the eye, without affecting the systemic absortion of topically applied opthalmic drugs.

• Archives of Pharmacal Research
• /
• v.26 no.1
• /
• pp.70-75
• /
• 2003

The objective of the present study was to investigate the uptake process of 4-Phenylazobenzoxycarbonyl-Pro-Leu-Gly-Pro-D-Arg (Pz-peptide), a hydrophilic and collagenase-labile pentapeptide, by isolated hepatocytes. For comparison, the uptake of Pz-peptide by Caco-2 cells and colonic cells, two known paracellular routes of Pz-peptide, was also evaluated. A simple and sensitive reversed-phase HPLC assay method using UV detection has been developed. The coefficient of variation for all the criteria of validation were less than 15%. The method was, therefore, considered to be sutable for measuring the concentration of Pz-peptide in the biological cells. Pz-peptide was extensively uptaked into hepatocytes. The initial velocity of Pz-peptide uptake assessed from the initial slope of the curve was plotted as Eadie-Hofstee plots. The maximum velocity ($V_{max}$) and the Michaelis constant ($K_m$) were 0.190$\pm$0.020 $nmol/min/10^6$ cells and 12.1$\pm$3.23 $\mu$M, respectively. The permeability-surface area product ($PS{influx}$) was calculated to be 0.0157 ml/min/10^6$ cells. $V_{max}$ and $K_m$ values for Caco-2 cells were calculated to be 6.22$\pm$0.930 pmol/min/10^6$ cells and 82.8$\pm$8.37 $\mu$M, respectively, being comparable with those of colonocytes (6.04$\pm$1.03 pmol/min/10^6$ cells and 87.8$\pm$13.2 $\mu$M, respectively). $PS_{influx}$ values for Caco-2 cells and colonocytes were calculated to be 0.0751 $\mu$l/min/10^6$ cells and 0.0688 $\mu$l/min/10^6$ cells, respectively. The more pronounced uptake of Pz-peptide by hepatocytes, when compared with Caco-2 cells and colonocytes, is probably due to its specific transporter. In conclusion, Pz-peptide, a paracellularly transported pentapeptide in the intestine and ocular epithelia, was uptaked into hepatocytes extensively. Although Pz-peptide is able to be uptaked into the Caco-2 cells and colonocytes, it is less pronounced when compared with hepatocytes. $PS_{influx}$ values of Caco-2 cells and colonocytes for unbound Pz-peptide under linear conditions were less than 0.4% when compared with that of hepatocytes.

• Archives of Pharmacal Research
• /
• v.19 no.3
• /
• pp.201-206
• /
• 1996

Nitric oxide (NO) is thought to be a second messenger involved in secretion. Upon stimulating pancreatic acinar cells with cholecystokinin-pancreozymin (CCK-PZ), NO formation has been shown to be associated with increased levels of cGMP (Seo et al., 1995). To elucidate the signaling pathway of VIP-induced enzyme secretion, we investigated the NO and cGMP synthesis steps as potential steps where two signal pathways triggered by CCK-PZ and VIP interact. The results obtained in this work provide evidence that increase in pancreatic enzyme secretion by treatment with VIP has no relationship with NOS activity and cGMP level. This conclusion was derived from the following findings that VIP treatment of rat pancreatic tissue increased amylase release as well as protein output in a dose- and time-dependent manner, whereas NOS activity and cGMP synthesis were not affected by VIP treatment as monitored by NOS activity assay and determining cGMP level, which was further confirmed by a NOS-inhibitor study. Consequently, CCK-PZ or VIP increases enzyme secretion in rat pancreatic tissue, but the two hormones are different in their mode of action. Together the results suggest that signaling pathway of VIP-induced enzyme secretion might either bypass the NO and cGMP synthesis steps or lie on a distinct pathway from CCK-PZ-induced pathway.

• The Korean Journal of Pharmacology
• /
• v.10 no.1 s.15
• /
• pp.47-52
• /
• 1974

The isolated rabbit gall bladder strips were prepared according to the technique described by Amer and Becvar (1969). The strips were placed in a bath containing 100 ml of Locke-Ringer solution maintained at $38^{\circ}C$. Oxygen was continuously bubbled through the solution. The tension of the muscle strip was initially adjusted to 0.7g. The contractile response was measured isometrically by a force-displacement transducer connected to a polygraph. The effect of a number of autonomic drugs were studied for their interaction with caerulein (Prof. V. Erspamer, F.I. 6934 Caerulein, Farmitalia, Italia), a gastrin or CCK.PZ like peptide, on isolated rabbit gall bladder strips. In this preparation caerulein produced contractions of CCK-PZ type, but the relative potency on a weight basis was 40 times that of CCK-PZ. The response of caerulein was not modified by either cholinergic or alpha or beta adrenergic blockade. However, the response of caerulein and of barium on the strips were prevented by papaverine or aminophylline. Isoproterenol, papaverine or aminophylline alone relaxed the preparation whereas caerulein, CCK-PZ, acetylcholine, serotonin, histamine or barium chloride contracted the preparation. In summary, it is concluded that caerulein on the gall bladder strip seems to act independently of the autonomic nervous system and mediated via mechanisms apparently similar to those involved in the action of barium chloride.

• The Korean Journal of Pharmacology
• /
• v.9 no.2
• /
• pp.17-27
• /
• 1973

Modifying the technique described by Schmidt, et al. (1972) the duodenum and stomach of female rats were perfused separately and contiunously with saline solution under urethane anesthesia. Secretory response of caerulein (Prof. V. Erspamer, F.I. 6934 Caerulein, Farmitalia, Italia), a gastrin or CCK-PZ like peptide, on acid, pepsin, bicarbonate and amylase were studied with and without simultaneous administration of secretin, CCK-PZ or other agents known secretory suppressives. A significant increase of acid, pepsin and amylase output was induced by intravenous infusion of caerulein. The response of acid secretion by caerulein in doses of 140 ng/100g/hr was equivalent to the response of histamine in the doses of $280\;{\mu}g/100g/hr$ and on a weight basis the potency of caerulein was approximately 2,000 times greater than histamine in rats. Acid secretory response of caerulein in the doses of 140 ng/100 g/hr was inhibited by simultaneous infusion of secretin in the doses of 0.2 u/ 100 g/hr, and the acid response was partly inhibited by concomitant infusion of histamine in the doses of $280\;{\mu}g/100g/hr$, but the response was enhanced by infusion of CCK-PZ in the doses of 0.2 u/100 g/hr. The secretory response of both aicd and enzymes were inhibited following administration of atropine in doses of 0.2 mg/100 g, but the response were not affected by hexamethonium in doses of 0.5 mg/100 g. In summary, it is concluded that caerolein is every effective in an increase of acid, pepsin and amylase secretion in rats through, possibly in part, the muscarinic and/or histaminic mechanism(s).