Targeted deletion of Hand2 in enteric neural precursor cells affects its functions in neurogenesis, neurotransmitter specification and gangliogenesis, causing functional aganglionosis.

Targeted deletion of the bHLH DNA-binding protein Hand2 in the neural crest, impacts development of the enteric nervous system (ENS), possibly by regulating the transition from neural precursor cell to neuron. We tested this hypothesis by targeting Hand2 deletion in nestin-expressing neural precursor (NEP) cells. The mutant mice showed abnormal ENS development, resulting in lethal neurogenic pseudo-obstruction. Neurogenesis of neurons derived from NEP cells identified a second nestin non-expressing neural precursor (NNEP) cell in the ENS. There was substantial compensation for the loss of neurons derived from the NEP pool by the NNEP pool but this was insufficient to abrogate the negative impact of Hand2 deletion. Hand2-mediated regulation of proliferation affected both neural precursor and neuron numbers. Differentiation of glial cells derived from the NEP cells was significantly decreased with no compensation from the NNEP pool of cells. Our data indicate differential developmental potential of NEPs and NNEPs; NNEPs preferentially differentiate as neurons, whereas NEPs give rise to both neurons and glial cells. Deletion of Hand2 also resulted in complete loss of NOS and VIP and a significant decrease in expression of choline acetyltransferase and calretinin, demonstrating a role for Hand2 in neurotransmitter specification and/or expression. Loss of Hand2 resulted in a marked disruption of the developing neural network, exemplified by lack of a myenteric plexus and extensive overgrowth of fibers. Thus, Hand2 is essential for neurogenesis, neurotransmitter specification and neural network patterning in the developing ENS.

Fetal head anomaly restricted to the eye, the mandible, and the pterygoid process of the sphenoid: a histological study.

A report on an unusual combination of anomalies in the head of a female fetus. The authors examined whole body semiserial paraffin sections of a female fetus (155 mm CRL; ∼18 weeks of gestation), with a particular focus on the head region. Cranial autonomic ganglia, nasal olfactory cells, and the orbital muscle were investigated using immunohistochemistry for tyrosine hydroxylase, vasoactive intestinal peptide, calretinin, and smooth muscle actin expression. The surface gross anatomy of the fetus appeared normal. The left eyeball lacked a lens (the eyeballs were otherwise normal). The orbital muscle was very thick and located in the anterolateral side of the extraocular muscles. Conversely, the extraocular muscles made a cluster in the superoposterior side of the orbit. The infratemporal fossa was small due to the bulky, transversely extended lateral pterygoid process in contrast to the small coronoid process of the mandible. The bilateral mandibular bases overlapped at the midline symphysis. The thin orbitosphenoid and thick alisphenoid provided an almost flat, anterior cranial base. Nasal olfactory cells and cranial autonomic ganglia appeared to be normal. No major anomaly was observed in the brain. Because of the changes in topographical anatomy, the orbital muscle probably lost its normal bony attachment and appeared to push the extraocular muscles superoposteriorly. A gene function redundancy rather than mutation may explain the present restricted anomalies in the mandible and pterygoid process.

Defects in sensory and autonomic ganglia and absence of locus coeruleus in mice deficient for the homeobox gene Phox2a.

Phox2a is a vertebrate homeodomain protein expressed in subsets of differentiating neurons. Here, we show that it is essential for proper development of the locus coeruleus, a subset of sympathetic and parasympathetic ganglia and the VIIth, IXth, and Xth cranial sensory ganglia. In the sensory ganglia, we have identified two differentiation blocks in Phox2a-/- mice. First, the transient expression of dopamine-beta-hydroxylase in neuroblasts is abolished, providing evidence that Phox2a controls noradrenergic traits in vivo. Second, the expression of the GDNF receptor subunit Ret is dramatically reduced, and there is a massive increase in apoptosis of ganglion cells, which are known to depend on GDNF in vivo. Therefore, Phox2a appears to regulate conventional differentiation traits and the ability of neurons to respond to essential survival factors.

Developmental genetics of neuroblastoma.

Case reports of neuroblastoma revealed that some individuals are genetically predisposed and that this genetic predisposition may have other consequences. According to a mutation model, two classes of individuals could acquire neuroblastoma. One (prezygotic) was a rare class that carried a dominant gene imparting high risk of the tumor. The other (postzygotic) comprised all other individuals, each at low risk. The model related tumor incidence to germinal and somatic mutation rates and thereby carried implications for environmental modification of tumorigenesis and demographic variation in incidence. Case reports also revealed associations of neuroblastoma with congenital defects and a susceptibility to second tumors. Analogy with retinoblastoma and Wilms' tumor of the kidney suggested that these associations could result from action of a neuroblastoma gene or from chromosomal aberration. One known dominantly inherited condition, von Recklinghausen's disease, could dispose to neuroblastoma and create some associations. According to the two-mutation model, neuroblastoma may have been a single recessive gene disorder at the level of the cell. The phenomena of aganglionosis, neuroblastoma in situ, maturation of neuroblastoma to ganglioneuroma, and spontaneous regression suggested that such a neuroblastoma gene interfered with normal developmental processes. The specificities of this gene and of those for von Recklinghausen's disease and pheochromocytoma suggested that the functiof a membrane macromolecule.

Rectal myectomy in Hirschsprung disease: a decade of experience.

Short-segment aganglionosis is becoming more frequently recognized in view of more sophisticated diagnostic techniques, as well as the greater clinical awareness of the entity. A series of 37 patients who underwent rectal myectomy during a ten-year period was reviewed. Selected patients with short-segment aganglionosis, proved by muscle biopsy, responded well to a single transanal operation that required an overnight hospital stay. The complications were minimal, and the cure or marked improvement has persisted for as long as 11 years. When major surgery of an abdominoperineal pull-through type was required after the myectomy (four patients), no technical difficulties were encountered. Within the limits of its applicability, rectal myectomy has a definite place in the treatment of short-segment Hirschsprung disease.

Modified Duhamel procedure for treatment of total aganglionic colon in childhood.

Based on this series of six patients with aganglionosis of the entire colon we conclude: (1) Radiographic findings of a shortened colon of normal caliber or the presence of "jejunalization" of the colon suggest total colonic aganglionosis in patients with a suggestive history. (2) All infants with persistent obstipation, distention, and poor weight gain should have a punch biopsy of the rectum even if the barium enema is normal. (3) The Martin modification of Duhamel's operation gives functional results comparable to those achievable in children with short-segment Hirschsprung's disease. (4) The use of a stapling device to divide the septum between aganglionic colon and pulled-through ileum is less satisfactory than using crushing clamps.

Constipation and congenital disorders of the myenteric plexus.

Full-thickness muscle biopsies have been taken from patients with severe disabling chronic constipation that has not responded to conservative measures. Assessment by neurohistochemical techniques has revealed that a range of neuronal dysplasias of the myenteric plexus are responsible in many cases; these include aganglionosis (Hirschsprung's disease), hypoganglionosis and hyperganglionosis. In cases considered unlikely to be Hirschsprung's disease on clinical grounds, the procedure used has often been anorectal myectomy; this has not only provided tissue for diagnosis but has also been of therapeutic value in most cases of hypoganglionosis and some cases reported as 'normal'.

[Follow-up and evolution of 10 cases operated on for total colonic aganglionosis]. / Seguimiento y evolucion de 10 casos intervenidos de aganglionismo colico total.

One hundred sixty-one cases of HIRSCHSPRUNG's disease have been operated on by us between 1966 and 1988. Of these, 19 were total colonic aganglionosis. Of eleven that underwent surgery before 1982, five are still alive. The eight that were operated on after 1982 have a good evolution. This report is a retrospective study of the clinical evolution in ten of the thirteen survivors. The ages are comprised between 21 and 2 1/2 years. We have reviewed their general condition, weight, height, tolerance to oral intake, stool frequency, general laboratory determinations, circulating iron, transferrin, transferrin saturation, absorption of vitamin B12 and folic acid, hydrogen test and stool examination. Except for one case, the others are in a perfect nutritional and growth condition. DUHAMEL's technique was employed in eight cases and REHBEIN's technique, in two. Given our good results, we think that LESTER-MARTIN's technique is unnecessary, except for that cases with extensive small bowel involvement due to the serious problems of absorption that this represents.

Segmental dilatation of sigmoid colon in a neonate: atypical presentation and histology.

Segmental dilatation of the colon is a rare disorder of colonic motility in children, often presenting with severe constipation in older infants, children, and occasionally adults. It may mimic the commoner Hirschsprung disease clinicoradiologically but differs in that the ganglion cell morphology and distribution are typically normal in the colon. We report a neonate with segmental dilatation of the sigmoid colon who had an atypical clinical presentation and describe certain abnormalities in bowel histology (hypertrophied muscularis propria, nerve plexus, and ganglion cells located within the circular layer rather than the normal myenteric location), for the first time in the English literature.

Hes1 and Hes5 regulate the development of the cranial and spinal nerve systems.

The basic helix-loop-helix genes Hes1 and Hes5, known Notch effectors, regulate the maintenance of neural stem cells and the development of the central nervous system (CNS). In the absence of Hes1 and Hes5, the size, shape and cytoarchitecture of the CNS are severely disorganized, but the development of the peripheral nervous system remains to be analyzed. Here, we found that in Hes1;Hes5 double-mutant mice, the cranial and spinal nerve systems are also severely disorganized. In these mutant mice, axonal projections from the mesencephalic neurons to the trigeminal (V) ganglion become aberrant and the proximal parts of the glossopharyngeal (IX) and vagus (X) nerves are fused. The hypoglossal (XII) nerve is also formed poorly. Furthermore, the dorsal root ganglia are fused with the spinal cord, and the dorsal and ventral roots of the spinal nerves are lacking in many segments. These results indicate that Hes1 and Hes5 play an important role in the formation of the cranial and spinal nerve systems.

Total intestinal aganglionosis. An autosomal recessive condition?

A case of total intestinal aganglionosis in a sib of a previously recorded patient is presented. The number of cases now reported is 9 in six families. The possibility that this condition is a distinct entity inherited in an autosomal recessive manner is discussed.

Clinical, laboratory and prognostic features of congenital large intestinal motor dysfunction (pseudo-Hirschsprung's disease).

One hundred and forty-eight cases of congenital large intestinal motor dysfunction (pseudo-Hirschsprung's disease) were reported by members of the Japanese Society of Pediatric Surgeons during the past 20 years. The disorder was defined as a congenital, non-mechanical obstruction of the intestine with the presence of intramural ganglia in the terminal rectum. Intramural ganglia were abnormal in 77 cases, normal in 42, and could not be determined in 29. Of those with abnormal intramural ganglia, 54 had immature ganglia or hypoganglionosis (oligoganglionosis), 15 had neuronal intestinal dysplasia, and eight had a segmental anomaly. Of those with a normal myenteric plexus, 22 had chronic and twelve had suspected idiopathic intestinal pseudo-obstruction syndrome; eight had megacystis-microcolon-intestinal hypoperistalsis syndrome. While cases with both hypoganglionosis and normal intramural ganglia had normal acetylcholine esterase activity, a significantly greater number of patients with hypoganglionosis lacked normal rectoanal reflexes. Patients with hypoganglionosis, chronic idiopathic intestinal pseudo-obstruction syndrome, and megalocystis-microcolon-intestinal hypoperistalsis syndrome had poor prognoses with an overall mortality of 36.9%. These findings indicate that congenital large intestinal motor dysfunction remains a serious disease of childhood.

Association of megacolon with a new dominant spotting gene (Dom) in the mouse.

A new semidominant mutation in the mouse is described. In heterozygotes it produces white spotting and a deficiency of myenteric ganglion cells in the colon and, in homozygotes it is lethal prior to 13 days of gestation. The mutation, called dominant megacolon, symbol Dom, is located on chromosome (chr) 15, 20.6 +/- 1.6 units proximal to Ca. Hairy ears, Eh, a semidominant gene also on chr 15 is shown to have a suppressing effect on crossing over in this section of chr 15.

GDNF family receptors in the embryonic and postnatal rat heart and reduced cholinergic innervation in mice hearts lacking ret or GFRalpha2.

Members of the GDNF family, which are important during peripheral nervous system development and kidney organogenesis, signal via Ret and GFRalpha receptors. Here we have studied their possible role in heart development. Gfra1 was expressed in the endocardial cushion mesenchyme at E12 and later, in the developing and mature valves, and in the walls of the aorta and the pulmonary trunk. Gfra2 was expressed in the outer layers of the aorta and pulmonary trunk and in the valves at E18-P60. Endocardial cells showed moderate Gfra2 mRNA and protein expression between E12 and E15. Gfra3 mRNA was detected, mainly postnatally, in scattered cells of the atria and the great vessels. In embryonic and postnatal rat cardiac ganglia, Ret and Gfra2 transcripts were seen in the neurons, whereas Gfra1 and Gfra3 mRNA were preferentially found in non-neuronal cells within the ganglia. GFRalpha2 immunoreactivity was seen in both cardiac ganglion neurons and their nerve fibers. There were no obvious non-neuronal defects in hearts of Ret-, GFRalpha1-, or GFRalpha2-deficient mice, suggesting that these receptors are not essential for gross cardiac development. However, E18 Ret-deficient mice exhibited a reduced volume of cardiac ganglia and cholinergic innervation of the ventricular conduction system. Moreover, adult Gfra2(-/-) mice showed reduced cholinergic innervation by 40% in their ventricles and by 60% in the ventricular conduction system. These findings indicate that GFRalpha2/Ret signaling is required for normal cholinergic innervation of heart.

Acetylcholine-related bowel dysmotility in homozygous mutant NCX/HOX11L.1-deficient (NCX-/-) mice-evidence that acetylcholine is implicated in causing intestinal neuronal dysplasia.

BACKGROUND/PURPOSE: Homozygous mutant Ncx/Hox11L.1-deficient (Ncx-/-) mice develop mega-ileo-ceco-colon (mega-ICC) with a caliber change in the proximal colon. The authors investigated the mechanism of intestinal dysmotility in these mice. METHODS: Five-week-old Ncx-/- mice with mega ICC were compared with age-matched BDF1 control mice. Jejunum, ileum, and colon were excised from all mice and 1.0-cm-long strips of each organ, each with a resting tension of 0.5g, were suspended in an organ bath filled with Tyrode's solution at 37 degrees C and bubbled with a mixture of 95% oxygen and 5% carbon dioxide. Contractile responses to acetylcholine chloride (ACh), histamine, serotonin, and barium chloride (BaCl2) were recorded isometrically. RESULTS: For ACh, Ncx-/- mice had decreased distal colon circular muscle contraction only at lower doses and decreased distal colon longitudinal muscle contraction for all doses compared with controls (P <.05 or P <.01). In the proximal colon, Ncx-/- mice had increased circular muscle contraction only at higher doses and decreased longitudinal muscle contraction only at lower doses compared with controls (P <.01 or P <.05). ACh did not affect jejunum, and there were no significant effects on ileum. There was no response to histamine and serotonin by any part of the bowel, and the response to BaCl2 was the same for both Ncx-/- mice and controls. CONCLUSIONS: Only ACh differentially affected muscle contraction in Ncx-/- mice in the proximal and distal colon. Thus, ACh is implicated in causing the bowel dysmotility seen in Ncx-/- mice and human IND.