Kym C. Tolman

Desipramine modulation of σ and opioid peptide receptor expression in glial cells

Abstract Exposure of C 6 glial cell cultures to desipramine induced the appearance of opioid receptors and up-regulated σ receptors. Opioid binding was demonstrated with 3 H-etorphine and 3 H-dihydromorphine (DHM), but was not observed with the μ, δ and κ ligands 3 H-DAMGE, 3 H-DADLE or 3 H-(−)ethylketocyclazocine in the presence of specific blockers, respectively. Competition experiments with 3 H-DHM and either (−)naloxone or (+)naloxone indicated the presence of authentic opioid receptors. In similar studies with β-endorphin, its truncated form (1–27) or their N-acetyl derivatives, β-endorphin proved to have the highest affinity. Opioid receptors in glial cell aggregates were primarily κ, with few μ and δ sites. Desipramine increased B max values for κ but not μ and δ.

In Vitro and In Vivo Expression of Opioid and σ Receptors in Rat C6 Glioma and Mouse N18TG2 Neuroblastoma Cells

Abstract: Mouse N18TG2 neuroblastoma and rat C6 glioma cell lines were injected into male nude mice, and the tumors were passaged serially. At each generation, tumors were analyzed for δ opioid binding using [3H][d‐Ala2,d‐Leu5]enkephalin and for σ1 and σ2 binding with 1,3‐[3H]di‐o‐tolylguanidine in the presence and absence of 1 µM pentazocine. Receptor density (Bmax) and affinity (KD) were estimated by homologous competition binding assays. Opioid and σBmax values in the solid tumors were significantly lower than their original levels in vitro. KD values for opioid/σ ligands were similar in vitro and in vivo. With successive passages in the murine host, δ opioid and σ1 binding of the neuroblastoma‐derived solid tumors became undetectable. In contrast, σ2 receptor Bmax values were unchanged with successive passages of the neuroblastoma‐derived tumors and doubled in the nude mouse‐borne gliomas. When neuroblastoma‐derived solid tumors that were devoid of δ opioid binding were returned to culture, opioid receptors appeared to be up‐regulated as compared with their original in vitro levels. Serial passaging of these recultured cells in vivo again resulted in a rapid decline in opioid receptor content. The opioid data are consistent with our prior findings on opioid binding diminution in human brain tumors. The pattern of change for σ binding was more complex, with the σ2 response in late passages of the glioma being reminiscent of the formerly observed increase in number of σ sites in transformed human meninges, kidney, and colon tissue.

Distribution and metabolism of doxorubicin in rats undergoing testicular circulatory isolation

Several hundred thousand men receive chemotherapy each year; many are sterilized by this treatment. Temporary testicular circulatory isolation (TCI), a regional drug delivery approach to circumvent this, decreases doxorubicin-induced testicular injury in the rat and provides partial protection from doxorubicin-related infertility. We evaluated the distribution of doxorubicin and its metabolites (doxorubicinol and doxorubicin aglycone) in rats treated with TCI. In each of 56 male Sprague-Dawley rats, the left spermatic cord and gubernaculum were mechanically clamped for 45 minutes. Immediately after clamp application, these rats received doxorubicin (6 mg/kg, intravenous bolus) and were killed at seven time points after doxorubicin administration, ranging from 30 minutes to 48 hours. Twenty-one control rats were treated identically but did not receive TCI. Doxorubicin and its metabolites were extracted from tissue (left testis, right testis, left kidney, heart, left lung, liver) and serum and analyzed by high-performance liquid chromatography. In the TCI group, the distribution of the parent drug and doxorubicinol in tissue and serum closely approximated levels from doxorubicin-treated controls not receiving TCI in all organs except left testis. No anthracycline was detected at any time point in the left testis of the TCI group. These results indicate that TCI completely protects the testis from doxorubicin exposure in this model and that TCI does not affect distribution of doxorubicin in other organs.

Testicular cytotoxicity of intravenous procarbazine in rats

Although the testicular cytotoxicity of procarbazine has been evaluated in the rat, previous models have utilized routes other than the intravenous one, and have generally employed multiple-dose regimens. In this report, we describe testicular toxicity in the Sprague-Dawley rat following a single intravenous bolus of procarbazine (0-700 mg kg body weight), with necropsy 59 +/- 2 days later. Testicular toxicity was evaluated qualitatively by histology and quantitatively by testicular weight, sperm head count, repopulation index and epididymal index. Effects of procarbazine on heart, lung, liver and kidney histology were evaluated qualitatively. Progressive dose-dependent testicular atrophy and oligospermia occurred at low and intermediate dosages of procarbazine. Marked testicular atrophy, oligospermia and germinal hypoplasia were observed at high dosages (500 and 700 mg kg-1 body weight). LD50 at day 59 for procarbazine appears to be approximately 600 mg kg-1 body weight using this regimen. This model will facilitate the study of techniques to avoid drug-induced testicular damage.

Preservation of fertility following doxorubicin administration in the rat

Several hundred thousand men receive chemotherapy each year; many are sterilized by this treatment. Testicular circulatory isolation (TCI), a regional drug exclusion approach to circumvent chemotherapy-related infertility, lessens doxorubicin-induced testicular injury in the rat. We evaluated the effect of TCI on doxorubicin-induced infertility in this study. Thirty-two eight-week-old male Sprague-Dawley rats were used. Eight rats received TCI for 45 min. Eight received doxorubicin (i.v. bolus) plus sham surgery. Eight received i.v. doxorubicin given immediately after institution of TCI. Eight controls received sham surgery alone. Mating studies began 2 months later. Six of the 8 males receiving TCI alone were fertile. In the doxorubicin-treated, sham-operated group, 0 of 7 animals were fertile. In the doxorubicin-treated group which also received TCI, 2 of 7 males were fertile. In the sham-operated group, all 8 rats were fertile. This is the first evidence that regional drug exclusion technique can improve fertility in this model.

Testicular circulatory isolation: Not a cause of immune-mediated testis injury in the rat

AbstractBackground: Testicular circulatory isolation (TCI), a regional drug exclusion approach designed to prevent chemotherapy-induced male infertility, can reduce testicular drug exposure and preserve fertility. The immunological sequelae of this surgical procedure were investigated. Methods: Forty Sprague-Dawley rats received unilateral TCI for 45 min and were killed at intervals of up to 43 days later. Testicular histology was evaluated qualitatively using hematoxylin and eosin stain, a direct immunofluorescent technique for detection of antigen-antibody complexes, and an indirect immunofluorescent technique to detect circulating antitestis antibodies. Results: No immune-mediated injury was evident up to 43 days after TCI. Conclusion: The current work, taken together with previously published data, indicate that TCI produces no immunological damage in the rat testis. Because TCI is well tolerated in humans, this work also supports the institution of human clinical trials of this technique in men about to receive fertility-threatening chemotherapy.

Testicular circulatory isolation prevents testicular exposure to doxorubicin in a rat model

Steady progress in the treatment of cancer has increased the proportion of longterm survivors, and delayed treatment toxicity such as infertility is becoming an important consideration. Despite many reports documenting the adverse effects of cytotoxic drugs on fertility in men [1], a practical method to avoid this is not currently available. We are investigating a mechanical approach to this problem, which we have termed testicular circulatory isolation (TCI). This consists of temporarily interrupting the blood supply to the testis during drug administration. In the rat, testicular injury due to doxorubicin is reduced by TCI [1]. TCI can partially protect male rats from drug-induced infertility [2], and no long-term testicular injury results from TCI [3]. We now report an investigation of the mechanism of this protective effect.