Prostatic Metaplasia of the Vagina and Uterine Cervix: An Androgen-associated Glandular Lesion of Surface Squamous Epithelium.

Prostatic-type differentiation in the lower female genital tract is encountered rarely and its causes and clinical associations are not well established. Within the vagina, reports to date have invariably described ectopic prostatic-type differentiation as restricted to the lamina propria. We recently encountered a patient receiving testosterone for gender dysphoria whose vaginectomy specimen showed a prostatic glandular proliferation within the surface epithelium. To elucidate its potential association with androgen exposure, we sought similar lesions, resected over a 26-year period, from patients with exogenous or endogenous androgen excess. Thirteen cases were identified, involving the vagina (n=12) and exocervix (n=1). The most common clinical context was gender dysphoria with long-term testosterone therapy; the lesion was present in 7 of 8 gender-dysphoric patients examined. Four other patients had congenital disorders of sexual development associated with endogenous androgen excess (congenital adrenal hyperplasia, 46,XY disorder of sexual development, and ovotesticular disorder of sexual development). Two had no known exposure to androgen excess. Immunohistochemically, glands stained for NKX3.1 (100% of cases), androgen receptor (100%), CK7 (92%), and prostate-specific antigen (69%). Follow-up (median duration, 11 mo) showed no masses or neoplasia. We propose the designation "androgen-associated prostatic metaplasia" for this form of prostate tissue with distinctive clinical, histologic and immunohistochemical features. It is novel and previously unrecognized within the vagina. It is strikingly prevalent among patients undergoing gender-affirming surgery, an increasingly common procedure. Recognition is important to distinguish it from other potentially neoplastic glandular lesions and facilitate accrual of more follow-up data to better understand its natural history.

Ciclosporin-induced accessory breast tissue: Dramatic improvement after dose adjustment.

Accessory breast tissue is a relatively common congenital condition in which abnormal accessory breast tissue is seen as a mass anywhere along the course of embryologic mammary streak in addition to the presence of normal breast tissue. Ciclosporin therapy has been associated with benign breast disease in women. However, to the best of our knowledge, there are no reported cases of accessory breast tissue growth associated with ciclosporin therapy and regression after adjusting the dose. A 48-year-old woman had renal transplantation in 2009 with her brother as the donor. Her transplant follow-up over eight years had been unremarkable. She presented to our transplant follow-up clinic with bilateral painful axillary masses approximately 3 cm × 3 cm in diameter, not attached to the skin or underlying structures with no skin changes and no lymphadenopathy. Breast examination did not reveal any abnormalities. Her ciclosporin levels over the previous three years ranged between 130 and 150 ng/mL. These levels were within the acceptable recommended level of 100-150 ng/mL at that time (currently reduced to 80-120 ng/ml). Ultrasound of both axilla showed well-defined hypoechoic smooth outline masses in both axillary regions 3 cm × 4 cm. Fine-needle aspiration showed lesions consisting of cohesive ductal cells. The findings were consistent with accessory breast tissue with no evidence of inflammatory infiltrate or malignant changes. Her ciclosporin dose was reduced with the subsequent follow-up visits levels ranging between 90 and 110 ng/mL. Clinical examination four months later showed dramatic reduction in the axillary masses on both sides. Ultrasound confirmed the regression in the size of both masses. We conclude that ciclosporin was probably responsible for the formation of accessory breast tissue and reduction in the dose of ciclosporin resulted in substantial reduction in the tissue size.

Morphological differences in BMP-2-induced ectopic bone between solid and crushed hyaluronan hydrogel templates.

The possibility to affect bone formation by using crushed versus solid hydrogels as carriers for bone morphogenetic protein 2 (BMP-2) was studied. Hydrogels, based on chemical crosslinking between hyaluronic acid and poly(vinyl alcohol) derivatives, were loaded with BMP-2 and hydroxyapatite. Crushed and solid forms of the gels were analyzed both in vitro via a release study using ¹²5I radioactive labeling of BMP-2, and in vivo in a subcutaneous ectopic bone model in rats. Dramatically different morphologies were observed for the ectopic bone formed in vivo in the two types of gels, even though virtually identical release profiles were observed in vitro. Solid hydrogels induced formation of a dense bone shell around non-degraded hydrogel, while crushed hydrogels demonstrated a uniform bone formation throughout the entire sample. These results suggest that by crushing the hydrogel, the construct's three-dimensional network becomes disrupted. This could expose unreacted functional groups, making the fragment's surfaces reactive and enable limited chemical fusion between the crushed hydrogel fragments, leading to similar in vitro release profiles. However, in vivo these interactions could be broken by enzymatic activity, creating a macroporous structure that allows easier cell infiltration, thus, facilitating bone formation.

Rosiglitazone inhibits bone regeneration and causes significant accumulation of fat at sites of new bone formation.

Thiazolidinediones (TZDs), peroxisome proliferator-activated receptor gamma activators, and insulin sensitizers represent drugs used to treat hyperglycemia in diabetic patients. Type 2 diabetes mellitus (T2DM) is associated with a twofold increase in fracture risk, and TZDs use increases this risk by an additional twofold. In this study, we analyzed the effect of systemic administration of the TZD rosiglitazone on new bone formation in two in vivo models of bone repair, a model of drilled bone defect regeneration (BDR) and distraction osteogenesis (DO) and a model of extended bone formation. Rosiglitazone significantly inhibited new endosteal bone formation in both models. This effect was correlated with a significant accumulation of fat cells, specifically at sites of bone regeneration. The diminished bone regeneration in the DO model in rosiglitazone-treated animals was associated with a significant decrease in cell proliferation measured by the number of cells expressing proliferating cell nuclear antigen and neovascularization measured by both the number of vascular sinusoids and the number of cells producing proangiogenic vascular endothelial growth factor at the DO site. In summary, rosiglitazone decreased new bone formation in both BDR and DO models of bone repair by mechanisms which include both intrinsic changes in mesenchymal stem cell proliferation and differentiation and changes in the local environment supporting angiogenesis and new bone formation. These studies suggest that bone regeneration may be significantly compromised in T2DM patients on TZD therapy.

Temporal and spatial CGRP innervation in recombinant human bone morphogenetic protein induced spinal fusion in rabbits.

STUDY DESIGN: Animal experiment using a rabbit posterolateral intertransverse process fusion model. OBJECTIVE: To explore the temporal and spatial distribution of sensory nerve fibers expressing calcitonin-gene related peptide (CGRP) during spinal fusion induced by recombinant human bone morphogenetic protein-4 and the role of the CGRP innervation in ectopic bone formation and remodeling. SUMMARY OF BACKGROUND DATA: Sensory neuropeptide CGRP involved in local bone turnover has been evidenced but its underlying mechanism is poorly understood. Knowledge in the CGRP innervation in ectopic bone induced by bone morphogenetic proteins can help us to understand its role in bone turnover. METHODS: Twenty-seven New Zealand white rabbits underwent single level posterolateral intertransverse process fusion of the lumbar vertebrae with implantation of porous poly-d,l-lactic acid blocks loaded with 1.25 microg recombinant human bone morphogenetic protein-4 solution. Animals were killed and the operated lumbar vertebrae were harvested for histomorphological evaluation at 3 days (n = 3), 1 week (n = 6), 3 weeks (n = 6), 7 weeks (n = 6), and 12 weeks (n = 6) following surgery, respectively. RESULTS: New cartilage presented at 1 week postimplantation adjacent to the implant, reached a peak volume at week 3 followed by a drop till week 12 after its ossification. Trabeculae-like woven bone structure presented at week 3. CGRP-positive nerve fibers regenerated already at 3 days postimplantation, reached its peak density at week 3. The CGRP-positive fibers presented both in fibrous tissues adjacent to proliferating cartilages and in bone marrow of newly formed trabecular bone. CONCLUSIONS: The observed spatial and temporal regeneration of CGRP-positive nerve fibers in ectopic bone formation suggested CGRP innervation is associated with ectopic osteogenesis.

A case of unusual course of adolescent menorrhagia: decidual cast as a side effect of treatment.

OBJECTIVE: To present an unusual side effect of therapy for adolescent menorrhagia, with commentary on management options. DESIGN: Case report. SETTING: Tertiary gynecologic center. PATIENT(S): A 13-year-old girl presenting with profuse uterine bleeding. INTERVENTION(S): Dysfunctional uterine bleeding was diagnosed, and pharmacologic therapy with a monophasic oral contraceptive (OC) pill was introduced. MAIN OUTCOME MEASURE(S): A large decidual cast occurred during treatment. RESULT(S): After finishing therapy with OC followed by three cycles of cyclic progestogen, the patient experienced regular menses. CONCLUSION(S): A large decidual cast may occur during treatment of adolescent menorrhagia with OCs; patients should be informed about this possibility, to reduce stress connected with the appearance of this side effect.

Altered glutamate receptor - transporter expression and spontaneous seizures in rats exposed to methylazoxymethanol in utero.

PURPOSE: Brain malformations are a common cause of intractable epilepsy and cognitive dysfunction in children. Prenatal exposure to the teratogen methylazoxymethanol (MAM) is a rodent model of brain malformation featuring loss of lamination, clusters of displaced hippocampal cells, and pharmaco-resistance to antiepileptic drugs. In a normotopic hippocampus, expression of postsynaptic glutamate receptors and the transporters regulating neurotransmitter reuptake are critical factors modulating excitation and synaptic communication. Alterations in this system can have profound effects on overall excitability, cognitive function, and seizure thresholds. METHODS: Immunohistochemical techniques were used to analyze the expression of N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5 methylisoxazole-4-proprionic acid (AMPA) receptor subunits in rats exposed to MAM in utero (25 mg/kg, intraperitoneal injection). We also examined the expression of several glutamate transporters (EAAC1, vGLUT1, and vGLUT2). A video-electroencephalographic (video-EEG) system was used for long-term monitoring of adult MAM-exposed rats. RESULTS: Heterotopic hippocampal neurons exhibited striking reductions in GluR1 and EAAC1 expression; vGlut2 expression was prominent in these regions. Spontaneous electrographic seizures were verified in two animals. CONCLUSIONS: We conclude that glutamate receptor subunit and transporter expression are altered in animals exposed to MAM in utero. Further studies in the MAM model may provide greater insight into the potential disruptions in excitatory synaptic neurotransmission that can occur in a malformed brain.

Determinants of drug brain uptake in a rat model of seizure-associated malformations of cortical development.

We examined the blood-brain barrier (BBB) function in methylazoxymethanol acetate (MAM)-treated rats, a model of human developmental brain malformations. We found aberrant vessels morphology and serum albumin leakage in the heterotopic (malformed) hippocampus; these changes were associated with a significant increase in endothelial P-glycoprotein (P-gp) expression. Seizures exacerbated BBB leakage and greatly augmented P-gp expression in vessels and additionally in perivascular/parenchymal astrocytes. The effects of seizures were observed to a much larger extent in malformed than in normal brain tissue. The intrinsic changes in BBB function in MAM-exposed rats were associated with increased blood-to-brain penetration of ondansetron, a P-gp substrate. However, a marked reduction in drug brain levels was provoked by seizures, and this effect was reversed by selective blockade of P-gp activity with tariquidar. Changes in BBB function may critically contribute to determine the brain uptake and distribution of P-gp substrates in epileptic tissue associated with developmental malformations.

Neuronal apoptosis and gray matter heterotopia in microcephaly produced by cytosine arabinoside in mice.

Primary microcephaly can be accompanied by numerous migration anomalies. This experiment was undertaken to examine the pathogenesis of gray matter heterotopia and microcephaly that is produced after administering cytosine arabinoside (Ara-C) to mice. Pregnant mice were intraperitoneally injected with Ara-C at 30 mg/kg body weight on days 13.5 and 14.5 of gestation, and then their offspring were examined. On embryonic day 15.5, in the ventricular zone of the cingulate cortex, the neuroepithelial cells lacked BrdU immunoreactivity. Nestin-immunoreactive radial glial fibers and calretinin-positive subplate fibers were disrupted. TUNEL reaction was remarkable throughout the cerebral hemisphere. Subcortical heterotopia in the cingulate cortex and subependymal nodular heterotopia in the dorsolateral part of the lateral ventricles became detectable by the first day after birth. Thirty-two days after birth, microcephaly was apparent; subcortical heterotopia was observed to have increased in size while it was still located in the frontal and cingulate cortices. This experiment demonstrated that Ara-C induces neuronal apoptosis throughout the cerebral hemisphere. The immunohistochemical characteristics in the gray matter heterotopia suggest that both the subcortical and the subependymal heterotopias were formed by neurons originally committed to the neocortex. We conclude that the gray matter heterotopia that accompanies the microcephaly was produced by a disturbance of radial, tangential, and interkinetic neuronal migrations due to the toxicity of Ara-C in the immature developing brain.

Type I interferon production by tertiary lymphoid tissue developing in response to 2,6,10,14-tetramethyl-pentadecane (pristane).

Lymphoid neogenesis is associated with antibody-mediated autoimmune diseases such as Sjogren's syndrome and rheumatoid arthritis. Although systemic lupus erythematosus is the prototypical B-cell-mediated autoimmune disease, the role of lymphoid neogenesis in its pathogenesis is unknown. Intraperitoneal injection of 2,6,10,14-tetramethyl-pentadecane (TMPD, pristane) or mineral oil causes lipogranuloma formation in mice, but only TMPD-treated mice develop lupus. We report that lipogranulomas are a form of lymphoid neogenesis. Immunoperoxidase staining of lipogranulomas revealed B cells, CD4(+) T cells, and dendritic cells and in some cases organization into T- and B-cell zones. Lipogranulomas also expressed the lymphoid chemokines CCL21, CCL19, CXCL13, CXCL12, and CCL22. Expression of the type I interferon (IFN-I)-inducible genes Mx1, IRF7, IP-10, and ISG-15 was greatly increased in TMPD- versus mineral oil-induced lipogranulomas. Dendritic cells from TMPD lipogranulomas underwent activation/maturation with high CD86 and interleukin-12 expression. Magnetic bead depletion of dendritic cells markedly diminished IFN-inducible gene (Mx1) expression. We conclude that TMPD-induced lupus is associated with the formation of ectopic lymphoid tissue containing activated dendritic cells producing IFN-I and interleukin-12. In view of the increased IFN-I production in systemic lupus erythematosus, these studies suggest that IFN-I from ectopic lymphoid tissue could play a role in the pathogenesis of experimental lupus in mice.

Embryonic and early postnatal abnormalities contributing to the development of hippocampal malformations in a rodent model of dysplasia.

While there are many recent examples of single gene deletions that lead to defects in cortical development, most human cases of cortical disorganization can be attributed to a combination of environmental and genetic factors. Elucidating the cellular or developmental basis of teratogenic exposures in experimental animals is an important approach to understanding how environmental insults at particular developmental junctures can lead to complex brain malformations. Rats with prenatal exposure to methylazoxymethanol (MAM) reproduce many anatomical features seen in epilepsy patients. Previous studies have shown that heterotopic clusters of neocortically derived neurons exhibit hyperexcitable firing activity and may be a source of heightened seizure susceptibility; however, the events that lead to the formation of these abnormal cell clusters is unclear. Here we used a panel of molecular markers and birthdating studies to show that in MAM-exposed rats the abnormal cell clusters (heterotopia) first appear postnatally in the hippocampus (P1-2) and that their appearance is preceded by a distinct sequence of perturbations in neocortical development: 1) disruption of the radial glial scaffolding with premature astroglial differentiation, and 2) thickening of the marginal zone with redistribution of Cajal-Retzius neurons to deeper layers. These initial events are followed by disruption of the cortical plate and appearance of subventricular zone nodules. Finally, we observed the erosion of neocortical subventricular zone nodules into the hippocampus around parturition followed by migration of nodules to hippocampus. We conclude that prenatal MAM exposure disrupts critical developmental processes and prenatal neocortical structures, ultimately resulting in neocortical disorganization and hippocampal malformations.

Ethanol induces heterotopias in organotypic cultures of rat cerebral cortex.

Abnormalities in the migration of cortical neurons to ectopic sites can be caused by prenatal exposure to ethanol. In extreme cases, cells migrate past the pial surface and form suprapial heterotopias or 'warts'. We used organotypic slice cultures from 17-day-old rat fetuses to examine structural and molecular changes that accompany wart formation. Cultures were exposed to ethanol (0, 200, 400 or 800 mg/dl) and maintained for 2-32 h. Fixed slices were sectioned and immunolabeled with antibodies directed against calretinin, reelin, nestin, GFAP, doublecortin, MAP-2 and NeuN. Ethanol promoted the widespread infiltration of the marginal zone (MZ) with neurons and the focal formation of warts. The appearance of warts is time- and concentration-dependent. Heterotopias comprised migrating neurons and were not detected in control slices. Warts were associated with breaches in the array of Cajal-Retzius cells and with translocation of reelin-immunoexpression from the MZ to the outer limit of the wart. Ethanol also altered the morphology of the radial glia. Thus, damage to the integrity of superficial cortex allows neurons to infiltrate the MZ, and if the pial-subpial glial barrier is also compromised these ectopic neurons can move beyond the normal cerebral limit to form a wart.

Radial and tangential neuronal migration disorder in ibotenate-induced cortical lesions in hamsters: immunohistochemical study of reelin, vimentin, and calretinin.

To investigate the mechanisms of radial and tangential neuronal migration disorders, immunohistochemical expressions of reelin, vimentin, and calretinin were examined in brain lesions induced by ibotenate (an agonist of the N-methyl-D-aspartate [NMDA] complex receptor) in hamsters. Thirty-four newborn hamsters were subjected to intracerebral injections of ibotenate, and 12 animals served as the control. These hamsters were examined at 1, 2, 3, 5, and 7 days after injections. The cortical lesions observed after ibotenate injections had a strong resemblance to the following neuronal migration disorders: (1) microgyria, (2) focal subcortical heterotopia, and (3) leptomeningeal glioneuronal heterotopia. In microgyria, the radial glial fibers were sparsely distributed, but in leptomeningeal glioneuronal heterotopia, vimentin-positive fibers extended into this abnormal neural tissue. Calretinin-immunoreactive neurons and fibers were present along the lesion forming the microgyria and abnormal neuronal arrangement. Focal subcortical heterotopia also included a small number of calretinin-expressing neurons originating from the subplate neuronal population. These results imply that the neuronal migration disorders produced by ibotenate show not only the migrational arrest of neurons but also interference from the termination of the migration process. We also suggest that the heterotopic neurons constituting the focal subcortical heterotopia originate in the lateral or medial ganglionic eminence of the ventral telencephalon, probably caused by the abnormal tangential neuronal migration.

Local and controlled release of growth factors (combination of IGF-I and TGF-beta I, and BMP-2 alone) from a polylactide coating of titanium implants does not lead to ectopic bone formation in sheep muscle.

The osteoinductive potential of growth factors leads not only to a stimulated bone formation in bony tissue but also in extra skeletal tissue. This potential depends on the dosage and potentially on the application method and may limit the clinical use. The aim of the present study was to investigate the potential of IGF-I, TGF-beta1 and BMP-2 released from a newly developed application systems of orthopaedic implants to induce ectopic bone formation in muscles. This bioactive coating showed a stimulating effect on fracture healing in several experimental studies before. Titanium discs were coated on one side with the drug carrier poly(d,l-lactide) (PDLLA), with the carrier plus IGF-I and TGF-beta1 or with the carrier plus BMP-2. The discs were implanted in the Musculus cleidomastoideus of sheep and followed up for 3 months. X-rays were taken after the operation and the day of sacrifice. The muscles plus implant were harvested and prepared for histology. Neither the radiology nor the histology revealed any signs of ectopic ossification in the implant/muscle interface or in a distance to the plate in any group. An influence of the locally applied growth factor, however, was seen in the formation of a soft tissue capsule. Histomorphometric analysis revealed a significantly larger capsule area over the growth factor coated side in comparison to the uncoated side or the pure titanium plate, indicating an effect of the applied growth factors on cells, however, not resulting in osteoinduction in muscle. The result showed that the local and controlled release of growth factors from PDLLA coated implants does not induce ectopic bone formation in sheep muscle and could be used in orthopaedic surgery to increase healing without the risk of ectopic bone formation in the surrounding soft tissue.

The genesis of epileptogenic cerebral heterotopia: clues from experimental models.

The pre-natal administration of methylazoxymethanol acetate (MAM) in rats is able to induce cerebral heterotopia that share striking similarities with those observed in human periventricular nodular heterotopia, a cerebral dysgenesis frequently associated with drug-resistant focal seizures. In the present study, we investigated the mode of neurogenesis in cerebral heterotopia of MAM-treated rats, by analyzing post-natal cytoarchitectural features and time of neurogenesis using bromodeoxyuridine immunocytochemistry. The cytoarchitectural analysis demonstrated the existence, in the early post-natal period, of white matter cellular bands in close anatomical relationship with the heterotopia, which most likely serve as a reservoir of young, migrating neurons for the newly forming heterotopia. The birth dating analysis demonstrated that the period of generation of neurons within the heterotopia and adjacent white matter bands, was extended in comparison to corticogenesis in normal rat brains. In addition, it demonstrated that the heterotopia were formed through a rather precise outside-in (for cortical and periventricular heterotopia) and dorso-ventral (for intra-hippocampal heterotopia) neurogenetic pattern. We hypothesize that the MAM-induced ablation of an early wave of cortical neurons is sufficient to alter per se the migration and differentiation of subsequently generated neurons, which in turn set the base for the formation of the different types of heterotopia. On this basis, we suggest a neurogenetic scheme for MAM-induced heterotopia that can also explain the origin and intrinsic epileptogenicity of periventricular nodular heterotopia in humans.

Effects of high fat fish oil and high fat corn oil diets on initiation of AOM-induced colonic aberrant crypt foci in male F344 rats.

Modulating effects of high fat fish oil (HFFO) and high fat corn oil (HFCO) diets on azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) were studied in male F344 rats following 8 weeks of dietary treatment. The incidence of AOM-induced ACF was significantly lower in the proximal colon of rats fed the HFFO diets compared with rats fed the HFCO diets. No differential effects were found on enzyme activities that are involved in metabolic activation and detoxification of AOM. Activities of hepatic P450 IAI and P450 IIBI and hepatic and feacal levels of lipid peroxidation were increased by feeding the HFFO diet. Hepatic GST activity and plasma levels of PGE(2) were significantly lower in rats fed the HFFO diets compared with those fed the HFCO diets. These observations demonstrate that HFFO diets with high levels of n-3 PUFAs are also protective against preneoplastic lesions in the early stages of chemically induced colon carcinogenesis. It seems unlikely from our results that the inhibitory effect of a HFFO diet can be attributed to an altered metabolic activation and detoxification of AOM. Other mechanisms such as oxidative stress or reduction of PGE(2) levels may play an important role in the anticarcinogenic effects of n-3 PUFAs.

The NMDA receptor complex is altered in an animal model of human cerebral heterotopia.

Double intraperitoneal injections of methylazoxymethanol (MAM) in pregnant rats induce developmental brain dysgenesis with nodular heterotopia similar to human periventricular nodular heterotopia (PNH) and composed of hyperexcitable neurons. Here we analyzed the NMDA receptor complex and associated proteins in the heterotopic neurons of 2- to 3-month-old MAM-treated rats by means of a combined immunocytochemical/molecular approach. Our data demonstrated a clear reduction of p286-active form of alphaCaMKII and a selective impairment of both the targeting and the CaMKII-dependent phosphorylation of NR2A/B subunits in the postsynaptic membranes of the MAM-induced heterotopia. The reduced NR2A/B immunofluorescence of the cellular membrane was not due to reduced expression since it was decreased only in postsynaptic fractions but not in the homogenate. NMDA-NR1 and AMPA-GluR2/3 subunits, as well as PSD-95 and total alphaCaMKII protein levels, were not affected in MAM-treated rats, thus revealing that the overall composition of the postsynaptic fraction was not altered. These data clearly suggest that the molecular organization of the NMDA/alphaCaMKII complex is selectively altered in the postsynaptic compartment of heterotopic neurons. This alteration can play a role in determining the hyperexcitability of brain heterotopia in MAM rats as well as in human patients affected by PNH.

Neurogenesis in cerebral heterotopia induced in rats by prenatal methylazoxymethanol treatment.

We have previously demonstrated that the antiproliferative agent methylazoxymethanol acetate (MAM) is able to induce in rats cerebral heterotopia that share striking similarities with those observed in human periventricular nodular heterotopia (PNH), a cerebral dysgenesis frequently observed in human patients affected by drug-resistant focal epilepsy. In this study, we investigated the time-course of neurogenesis in the cerebral heterotopia of MAM-treated rats, with the idea of understanding why PNH develop in human patients. For these goals, we analyzed the cytoarchitectural features, the time of neurogenesis and the cellular phenotype of the heterotopia, by means of BrdU immunocytochemistry and confocal immunofluorescence experiments. Our data demonstrate that the different types of heterotopia in MAM-treated rats are formed through the same altered neurogenetic process, which follows quite organized neurogenetic gradients. The MAM-induced ablation of an early wave of cortical neurons is sufficient to alter per se the migration and differentiation of subsequently generated neurons, which in turn set the base for the formation of the different heterotopic structures. The neurogenesis of MAM-induced heterotopia may explain the origin and intrinsic epileptogenicity of periventricular nodular heterotopia in human patients.

An examination of calcium current function on heterotopic neurons in hippocampal slices from rats exposed to methylazoxymethanol.

PURPOSE: To study voltage-dependent calcium currents (VDCCs) on hippocampal heterotopic neurons by using whole-cell patch-clamp techniques in brain slices prepared from methylaxozymethanol (MAM)-exposed rats. METHODS: Whole-cell voltage-clamp recordings were obtained from visually identified neurons in acute brain slices by using an infrared differential interference contrast (IR-DIC) video microscopy system. Heterotopic neurons were compared with normotopic pyramidal cells in hippocampal slices from MAM-exposed rats or CA1 pyramidal neurons in slices from controls. RESULTS: Heterotopic neurons expressed a prominent VDCC, which exhibited a peak current maximum around -30 mV (holding potential, -60 mV) and an inactivation time constant of 48.2 +/- 2.4 ms (n = 91). VDCC peak current and inactivation time constants were similar for normotopic (n = 92) and CA1 pyramidal cells (n = 40). Pharmacologic analysis of VDCC, on heterotopic, normotopic, and CA1 pyramidal cells, revealed an approximately 70% blockade of peak Ca2+ current with nifedipine and amiloride (L- and T-type channel blockers, respectively). Inhibition of VDCC, for all three cell types, also was similar when more specific Ca2+ channel antagonists were used [e.g., omega-conotoxin GVIA (N-type), omega-agatoxin KT (P/Q-type), and sFTX-3.3 (P-type)]. VDCC modulation by norepinephrine (NE) or adrenergic receptor-specific agonists [clonidine (alpha2), isoproterenol (beta), and phenylephrine (alpha1)] was similar for heterotopic and CA1 pyramidal cells. CONCLUSIONS: Heterotopic neurons do not appear to exhibit Ca2+ channel abnormalities that could contribute to the reported hyperexcitability associated with MAM-exposed rats.