Transcriptome analysis of the fungal pathogen Rosellinia necatrix during infection of a susceptible avocado rootstock identifies potential mechanisms of pathogenesis.

BACKGROUND: White root rot disease caused by Rosellinia necatrix is one of the most important threats affecting avocado productivity in tropical and subtropical climates. Control of this disease is complex and nowadays, lies in the use of physical and chemical methods, although none have proven to be fully effective. Detailed understanding of the molecular mechanisms underlying white root rot disease has the potential of aiding future developments in disease resistance and management. In this regard, this study used RNA-Seq technology to compare the transcriptomic profiles of R. necatrix during infection of susceptible avocado 'Dusa' roots with that obtained from the fungus cultured in rich medium. RESULTS: The transcriptomes from three biological replicates of R. necatrix colonizing avocado roots (RGA) and R. necatrix growing on potato dextrose agar media (RGPDA) were analyzed using Illumina sequencing. A total of 12,104 transcripts were obtained, among which 1937 were differentially expressed genes (DEG), 137 exclusively expressed in RGA and 160 in RGPDA. During the root infection process, genes involved in the production of fungal toxins, detoxification and transport of toxic compounds, hormone biosynthesis, gene silencing and plant cell wall degradation were overexpressed. Interestingly, 24 out of the 137 contigs expressed only during R. necatrix growth on avocado roots, were predicted as candidate effector proteins (CEP) with a probability above 60%. The PHI (Pathogen Host Interaction) database revealed that three of the R. necatrix CEP showed homology with previously annotated effectors, already proven experimentally via pathogen-host interaction. CONCLUSIONS: The analysis of the full-length transcriptome of R. necatrix during the infection process is suggesting that the success of this fungus to infect roots of diverse crops might be attributed to the production of different compounds which, singly or in combination, interfere with defense or signaling mechanisms shared among distinct plant families. The transcriptome analysis of R. necatrix during the infection process provides useful information and facilitates further research to a more in -depth understanding of the biology and virulence of this emergent pathogen. In turn, this will make possible to evolve novel strategies for white root rot management in avocado.

Gastric tumor from metastasis of breast cancer.

Metastatic tumours of the stomach have been reported to result from various types of cancer. Among them, gastric metastasis from breast cancer has been recognised in 0.3-18% patients (1-4). Here, a rare case of metastatic gastric tumour derived from breast carcinoma is reported. Gastric endoscopy confirmed a large, friable mass (approximately 5 cm in diameter) in the upper part of the gastric body. The mass within the stomach was difficult to distinguish from primary gastric cancer, although biopsies of this lesion revealed the characteristics of adenocarcinoma. In addition, immunohistochemistry showed the positive expression of mammaglobin. Taken together, the evidence pointed to metastasis of breast cancer to the stomach. The patient was treated with hormonal therapy (letrozole), and the size of the metastasis in the stomach was markedly reduced. Therefore, a gastric metastasis from breast cancer was diagnosed successfully using immunohistochemistry and unnecessary surgery was avoided. In conclusion, although gastric metastatic tumours derived from breast carcinoma are rare, their accurate pre-operative diagnosis and appropriate systemic treatment is essential.

GFP sheds light on the infection process of avocado roots by Rosellinia necatrix.

In order to monitor Rosellinia necatrix infection of avocado roots, we generated a plasmid vector (pCPXHY1eGFP) constitutively expressing EGFP and developed a protoplast transformation protocol. Using this protocol, four R. necatrix isolates were efficiently transformed and were shown to stably express EGFP homogeneously while not having any observable effect on pathogenicity. Confocal laser scanning microscopy (CLSM) images of avocado roots infected with the highly virulent isolate CH53-GFP demonstrated that fungal penetration of avocado roots occurs simultaneously at several random sites, but it occurs preferentially in the crown region as well as throughout the lenticels and in the junctions between epidermal cells. Not only were R. necatrix hyphae observed invading the epidermal and cortical root cells, but they were also able to penetrate the primary and secondary xylem. Scanning electron microscopy (SEM) images allowed detailed visualisation of the hyphal network generated by invasion of R. necatrix through the epidermal, cortical and vascular cells, including hyphal anastomosis and branching points. To our knowledge, this is the first report describing the construction of GFP-tagged strains belonging to the genus Rosellinia for monitoring white root rot using CLSM and SEM.

Molecular diversity of ecologically distinct Mal de Río Cuarto virus isolates based on restriction fragment length polymorphism (RFLPs) and genome sequence analysis of segments 1, 7, 9 and 10.

Viruses of the species Mal de Río Cuarto virus (genus Fijivirus, family Reoviridae) cause significant economic losses in maize in Argentina. Genetic changes in the virus genome leading to better adaptation to diverse ecological conditions were postulated that would account for the increasing MRCV variability. The genomic differences between MRCV isolates from four ecologically different areas (Río Cuarto, RC; Pergamino, P; Jesús María, JM; and Tafí del Valle, TV) were studied. RT-PCR-amplified fragments comprising four genomic segments (Seg1, Seg7, Seg9 and Seg10) of MRCV isolates were compared by RFLPs and nucleotide sequences. The segments were chosen based on the proteins they encode: RNA-dependent-RNA polymerase, proteins putatively associated with tubular structures and viroplasm and the major outer capsid protein, respectively. Genetic comparison suggested that JM and TV isolates were genetically similar, but RC and P were different. Therefore, they were clustered in three genetic groups (JM = TV, RC and P). Together, nucleotide and amino acid sequence identities of the genomic segments were often above 96%. Seg1 was more variable (viral polymerase), whereas Seg7 (putative tubular structure) was the most conserved. Phylogeny analysis showed that MRCV isolates could be clustered in 'mountain area' and 'high production area' groups according to their geographical occurrence.

Evidence for involvement of glial cell activity in the control of extracellular D-serine contents in the rat brain.

The continuous intra-cortical infusion of a glia toxin, fluorocitrate, at the concentration of 1 mM caused a decrease in the cortical extracellular contents of an intrinsic coagonist for the N-methyl-D-aspartate (NMDA) type glutamate receptor, D-serine, by peaking at 40 min by -25% but produced an increase in those of glycine and L-serine. The attenuated glial activity by fluorocitrate was verified by a marked reduction in the extracellular glutamine contents. The present findings suggest that a group of glial cells such as a population of the protoplasmic astrocytes could, at least in part, participate differently in the regulation of the extracellular release of D-serine and another NMDA coagonist glycine in the medial frontal cortex of the rat.

Infection of Rosellinia necatrix with purified viral particles of a member of Partitiviridae (RnPV1-W8).

Isolate W8 of the white root rot fungus, Rosellinia necatrix, harbors three dsRNA segments, L1-, L2- and M-dsRNAs, and showed an irregular colony margin, slow growth, and moderate virulence. The M-dsRNA was previously shown to be the genome of a partitivirus, RnPV1-W8. Here a transfection protocol was developed for RnPV1-W8. Protoplasts of two virus-free isolates of R. necatrix were inoculated with purified viral particles using a polyethylene glycol-mediated method. Virus infection was confirmed by electrophoresis and Northern analysis. RnPV1-W8 introduced into the new host isolates was transmissible via hyphal anastomosis. However, the infection had no effect on the morphology and virulence of infected isolates of R. necatrix. This is the first report on the transfection of a partitivirus for R. necatrix.

A Reovirus Causes Hypovirulence of Rosellinia necatrix.

ABSTRACT White root rot, caused by Rosellinia necatrix, is a serious soilborne disease of fruit trees and other woody plants. R. necatrix isolate W370 contains 12 segments of double-stranded RNA (dsRNA) that is believed to represent a possible member of the family Reoviridae. W370 was weakly virulent and its hyphal-tip strains became dsRNA free and strongly virulent. The 12 segments of W370dsRNA were transmitted to hygromycin B-resistant strain RT37-1, derived from a dsRNA-free strain of W370 in all or none fashion through hyphal contact with W370. The W370dsRNA-transmitted strains were less virulent than their parent strain RT37-1 on apple seedlings, with mortality ranging between 0 to 16.7% in apple seedlings that were inoculated with the W370dsRNA-containing strains and 50 to 100% for seedlings inoculated with the dsRNA-free strains. Some W370dsRNA-containing strains killed greater than 16.7% of seedlings, but these were found to have lost the dsRNA in planta. These results indicate that W370dsRNA is a hypovirulence factor in R. necatrix. In addition, a strain lost one segment (S8) of W370dsRNA during subculture, and the S8-deficient mutant strain also exhibits hypovirulence in R. necatrix.

Characterization of the small subgenomic RNA of Soybean dwarf virus.

We have characterized a small subgenomic RNA of Japanese strains of Soybean dwarf virus (SbDV). Northern blot analyses of SbDV-infected plants showed that the small RNA contained the 3' terminal sequence of the genome and was detected in four typical Japanese SbDV strains, YS, YP, DS and DP. In the case of SbDV-DS, the RNA was 220 nucleotides in length and was transcribed from the 3' terminal region of the genome. This RNA appeared at a similar time to genomic RNA and a large sgRNA, and thereafter persisted in the infected plant. Since no conserved open reading frame (ORF) among the strains was postulated in the 3' terminal region, the small subgenomic RNA may have some regulatory roles in SbDV infections.

First Report of Tulip band breaking virus in Mosaic Diseased Tulip in Japan.

Tulip (Tulipa spp.) is an ornamental plant of major economic importance in Japan. Regions in Toyama Prefecture are some of the most productive for producing tulip bulbs, shipping approximately 50 million bulbs annually. However, mosaic diseases caused by viruses such as Tulip breaking virus (TBV) currently limit bulb production in these areas. Only the potyviruses TBV and Lily mottle virus (LMoV) have been reported infecting tulip in Japan. A virus isolate from tulip with flower-breaking symptom in Toyama Prefecture was tentatively named OE4 and was presumed to be LMoV after detection by LMoV-specific reverse transcription-polymerase chain reaction (RT-PCR) (4). When OE4 was mechanically inoculated on test plants (13 species from six families), RT-PCR confirmed that it infected plants in the Liliaceae (Tulipa spp., Lilium formosanum, and L. concolor) with mosaic symptoms but did not induce any symptoms in Chenopodium quinoa, Tetragonia tetragonoides, and Nicotiana benthamiana. According to Dekker et al. (2) LMoV and Tulip band breaking virus (TBBV) infected Tulipa spp. and TBBV did not infect C. quinoa, T. tetragonoides, N. clevelandii, and N. benthamiana, species that were local or systemic hosts for LMoV. To analyze the genomic sequence of OE4, a primer set was designed for amplifying the coat protein (CP) gene of LMoV, with 5' -GCAAATGAGACACTCAATG-3' as a forward primer and 5'-TTACATAGAAATTCCAAGTAAG-3' as a reverse primer. The fragment obtained was cloned and sequenced (DDBJ/EMBL/GenBank Accession No. AB078007). The CP gene of OE4 consisted of 825 nucleotides and had 86.5% identity (90.6% identity in deduced amino acid sequence) with the CP gene of LMoV-Netherlands (S44147) (3). When it was compared with partial sequences (277 nucleotides) of LMoV (S60810) and TBBV (S60805) (2), the nucleotide sequence identities were 89.9 and 96.4%, respectively. Multiple alignment and a phylogenetic tree based on the 277 nt of the tulip potyviruses, including OE4, showed the close relationship between OE4 and TBBV. These results indicated that OE4 was an isolate of TBBV. However, the CP amino acid sequence identity between TBBV and LMoV was more than 80%, and it seemed logical that TBBV be assigned to a strain of LMoV, according to potyvirus species demarcating criteria (1). Another screening using RT-PCR based detection, and an inoculation test for 55 flower-breaking tulips collected from fields in Toyama Prefecture revealed that four tulip plants were infected with TBBV and 54 with TBV. This suggests that TBV is more prevalent than TBBV, and we need to produce an attenuated virus of TBV, which will be effective for managing tulip breaking disease in the prefecture. References: (1) P. H. Berger et al. Family Potyviridae. Pages 703-724 in: Virus Taxonomy. Academic Press, San Diego, CA, 2000. (2) E. L. Dekker et al. J. Gen. Virol. 74:881, 1993. (3) S. A. Langeveld et al. J. Gen. Virol. 72:1531, 1991. (4) T. Se and S. Kanematsu. Ann. Phytopathol. Soc. Jpn. 64:420, 1998.

Vesicular prurigo pigmentosa cured by minocycline.

We present a case of prurigo pigmentosa associated with vesicles that we call 'vesicular prurigo pigmentosa'. The subject was treated using minocycline with good results and no recurrence of the lesions over a 2-year period.

Comparison of complete nucleotide sequences of genomic RNAs of four Soybean dwarf virus strains that differ in their vector specificity and symptom production.

Soybean dwarf virus (SbDV) is divided into four strains, namely YS, YP, DS and DP. YS and YP cause yellowing in soybeans, while DS and DP cause dwarfing. YS and DS are transmitted by Aulacorthum solani, while YP and DP are transmitted by Acyrthosiphon pisum. To clarify the taxonomic relationship between the four strains and to classify SbDV into an appropriate genus in the Luteoviridae, we determined the complete nucleotide sequences of genomic RNAs of four isolates belonging to each of the strains. The genomes of the four isolates had a chimeric form between Barley yellow dwarf virus-PAV and poleroviruses, and the genome organizations were similar to the Australian isolate SbDV Tas-1. In all of the non-coding regions and ORFs, nucleotide and deduced amino acid sequence identity between the same symptom-type strains was higher than that between the different symptom-type strains. However, in the N-terminal half of the readthrough domain (RTD) the deduced amino acid identity between the same aphid transmissibility-type strains was higher than that between the different aphid transmissibility-type strains. These data suggest that the N-terminal half of the RTD is closely related to the aphid transmission specificity, and that the present strains were generated from ancestral Y and D strains by mutations and strong selection pressures of efficient aphid transmission. Therefore, we propose that SbDV should be classified into a new genus in the family Luteoviridae and that the four strains described should be regarded as different strains of the same virus, rather than as distinct virus species.

The Japanese Study Group of Insulin Therapy for Childhood and Adolescent Diabetes (JSGIT): initial aims and impact of the family history of type 1 diabetes mellitus in Japanese children.

The Japanese Study Group of Insulin Therapy for Childhood and Adolescent Diabetes (JSGIT) was established in July 1994 with the chief aim to improve the quality of therapy for type 1 diabetes in children, an entity far less common in Japan than in Europe. We proposed four initial research topics: (i) to determine the current status of medical care and glycemic control in Japanese children with type 1 diabetes mellitus; (ii) to standardize the measurement of hemoglobin A1c; (iii) to establish a registry of a large cohort of patients in order to enable prospective studies to improve the quality of therapy for children with type 1 diabetes in Japan; and (iv) to enable participants of the JSGIT to hold a workshop twice annually. We registered a total of 736 patients from 45 hospitals throughout Japan. Intervention via insulin treatment was instituted after 2 yr for those patients whose hemoglobin A1c level was more than 8.1%. The proportion of patients receiving multiple insulin injections increased after intervention; however, average hemoglobin A1c in females remained significantly higher than in males. We identified two forms of diabetes in Japanese children: a rapidly progressive form and a more slowly progressive form. There was a significantly higher prevalence of a family history of diabetes in first-degree relatives in the slowly progressive form. These preliminary findings are the result of the first collaborative study of childhood diabetes in Japan.

Serum parathyroid hormone and calcitonin levels in racehorses with fracture.

Serum parathyroid hormone (PTH) and calcitonin (CT) levels in fractured racehorses were measured by radioimmunoassay. Racehorses with fracture of large bone such as the radius, third metacarpus, third carpus, digital bone or tibia, showed normal PTH level and elevated CT level in the serum. Serum PTH level was slightly higher in racehorses with sesamoid bone fracture compared to that of healthy racehorses, but not statistically significant. Moreover, serum CT level of racehorses with sesamoid bone fracture was significantly higher than that of healthy racehorses. Racehorses with sesamoid bone fracture and large bone fracture might be in different conditions of calcium regulation.

Infection Behavior of Venturia nashicola, the Cause of Scab on Asian Pears.

ABSTRACT The infection of Japanese pear by Venturia nashicola, the cause of scab on Asian pears (Japanese pear, Pyrus pylifolia var. culta; Chinese pear, P. ussuriensis), was examined using light and electron microscopy to determine the mechanism of resistance in pears. Early stages of infection were similar on the susceptible cv. Kosui, the resistant cv. Kinchaku, and the nonhost European pear (P. communis) cv. Flemish Beauty. V. nashicola penetrated only the cuticle layer on pear leaves and formed subcuticular hyphae on all three cultivars. Hyphae were localized in the pectin layer of pear leaves and never penetrated into the cytoplasm of epidermal cells. This restriction of fungal growth suggested that pectinases released by infection hyphae or subcuticular hyphae may be important in infection. Subcuticular hyphae were modified ultrastructurally in the pectin layer of resistant pear cultivars accompanied by fungal cell death. In contrast, fungal cells appeared intact in susceptible pear cultivars, suggesting the existence of resistance mechanisms.

Generation of superoxide anion and localization of CuZn-superoxide dismutase in the vascular tissue of spinach hypocotyls: their association with lignification.

The sites of generations of superoxide anions and hydrogen peroxide in cross sections of hypocotyls from spinach seedlings were located by staining with nitroblue tetrazolium (NBT) and with starch-iodide, respectively. Formazan, produced upon the reduction of NBT by superoxide, was observed mainly in the vascular tissue only in the presence of inhibitors of CuZn-superoxide dismutase (CuZn-SOD), and its formation was suppressed under anaerobic conditions. Thus, NBT was reduced to formazan specifically by the superoxide anions generated in vascular tissue. The reduction of NBT was suppressed by inhibitors of NAD(P)H oxidase, but neither by cyanide nor azide, indicating the involvement of NAD(P)H oxidase in the generation of superoxide anions in the vascular tissue. Starch-I2 complex also was formed in the vascular tissue, but not in the presence of either the CuZn-SOD inhibitor or the NAD(P)H oxidase inhibitor, indicating that the hydrogen peroxide is produced via the catalytic disproportionation with CuZn-SOD of the superoxide generated by NAD(P)H oxidase. Generations of superoxide anions and hydrogen peroxide in the vascular tissue were particularly apparent in the xylem and associated with the sites of distribution of CuZn-SOD as determined by an immunohistochemical method, and also with the location of lignin as determined by the phloroglucin-HCl reaction.

Mutations in the hepatocyte nuclear factor-1alpha/MODY3 gene in Japanese subjects with early- and late-onset NIDDM.

Recent studies have shown that mutations in the hepatocyte nuclear factor (HNF)-1alpha gene are the cause of maturity-onset diabetes of the young type 3 (MODY3). We have screened 193 unrelated Japanese subjects with NIDDM for mutations in this gene: 83 with early-onset NIDDM (diagnosis at <30 years of age) and 110 with late-onset NIDDM (diagnosis > or = 30 years of age). All of the members of the latter group also had at least one sibling with NIDDM. The 10 exons, flanking introns, and promoter region were amplified using polymerase chain reaction and were sequenced directly. Mutations were found in 7 of the 83 (8%) unrelated subjects with early-onset NIDDM. The mutations were each different and included four missense mutations (L12H, R131Q, K205Q, and R263C) and three frameshift mutations (P379fsdelCT, T392fsdelA, and L584S585fsinsTC). One of the 110 subjects with late-onset NIDDM was heterozygous for the missense mutation G191D. This subject, who was diagnosed with NIDDM at 64 years of age, also had a brother with NIDDM (age at diagnosis, 54 years) who carried the same mutation, suggesting that this mutation contributed to the development of NIDDM in these two siblings. None of these mutations were present in 50 unrelated subjects with normal glucose tolerance (100 normal chromosomes). Mutations in the HNF-1alpha gene occur in Japanese subjects with NIDDM and appear to be an important cause of early-onset NIDDM in this population. In addition, they are present in about 1% of subjects with late-onset NIDDM.

Effects of growth hormone (GH)-releasing hormone and its analogs on GH secretion from cultured adenohypophysial cells in cattle.

The effects of the human growth hormone (GH)-releasing hormone (hGRF(1-44)-NH2: hGRF-44), bovine GRF (bGRF(1-44)-NH2: bGRF-44), and their analogs (hGRF(1-29)-NH2, [D-Ala2]- hGRF(1-29)-NH2, bGRF(1-29)-NH2,[D-Ala2,Ala15]-bGRF(1-29)-NH2) as well as rat GRF(rGRF) on bovine GH release from anterior pituitary (AP) cells were studied in vitro in steers. The AP cells were incubated for 2 hr with the GRFs after preincubation for 3.5 d. Both of the hGRF-44 and hGRF(1-29)-NH2 (hGRF-29) significantly stimulated GH release from cultured cells at doses from 10(-14) to 10(-8)M (P < 0.01). The analog [D-Ala2]-hGRF-29 significantly induced GH release in media at doses from 10(-18) to 10(-8) M (P < 0.01). The bGRF-44, bGRF(1-29)-NH2 (bGRF-29), and [D-Ala2, Ala15]-bGRF-29 significantly induced GH release in media at doses as low as 10(-18), 10(-17), and 10(-16)M, respectively (P < 0.01). At doses from 10(-11) to 10(-8)M, there were no significant differences in GH-releasing potency between the hGRFs and bGRFs. The rGRF significantly stimulated GH release at doses ranging from 10(-14) to 10(-8) M (P < 0.01). The linear regression tests showed that the hGRFs, bGRFs, and rGRF, at doses from 10(-14) to 10(-8)M, induced GH release in a dose-related manner (P < 0.01). These results suggest that the hGRF, bGRF, and their analogs, as well as rGRF, are potent secretagogues of GH release from adenohypophysial cells in vitro in cattle.

Effects of atipamezole, an alpha 2-adrenergic antagonist, and somatostatin on xylazine-induced growth hormone release in calves.

In order to clarify the mechanism of xylazine-induced GH release, we investigated the effects of atipamezole, a selective alpha 2-adrenergic antagonist, and somatostatin (SRIF) on xylazine-stimulated GH release in calves. Xylazine injection (0.30 mg/kg BW, iv) induced a rapid increase in the GH concentration. When atipamezole was used in combination with xylazine, it blunted the increase in the plasma GH concentration induced by the xylazine injection. The GH levels at 15-50 min after the simultaneous injection of xylazine and atipamezole were significantly (P < 0.05) lower than the corresponding values in the animals given xylazine alone. The area under the GH response curve for 120 min after the simultaneous injection of xylazine and atipamezole was significantly (P < 0.05) smaller than that for the xylazine alone. A series of five intravenous injections of 1 mg of SRIF at 10-min intervals also blunted xylazine-stimulated GH release. Atipamezole partially suppressed xylazine-induced hyperglycemia, but SRIF completely suppressed the hyperglycemia for the first 60 min after the xylazine injection and the suppression by SRIF was stronger than that by atipamezole. On the other hand, both atipamezole and SRIF failed to blunt xylazine-induced hypoinsulinemia. The present results suggest that xylazine stimulates GH release via the alpha 2-adrenergic pathway in cattle, but the mechanism of xylazine-induced hyperglycemia remains to be determined.

The effects of xylazine on plasma concentrations of growth hormone, insulin-like growth factor-I, glucose and insulin in calves.

The purpose of the present study was to examine the responses of plasma growth hormone (GH) and insulin-like growth factor-I (IGF-I) levels to intravenous injection of xylazine in female dairy calves. Xylazine (0.05, 0.15 and 0.30 mg/kg body wt., i.v.) injections induced a significant dose-dependent increase in plasma GH level within 30 min. After plasma GH levels reached peaks, GH concentrations began to decrease immediately and they returned to control levels 1 h after xylazine injection. Plasma IGF-I concentration tended to be suppressed by xylazine treatment. Xylazine induced a significant dose-dependent increase in plasma glucose for 3.5 to 5.5 h after the treatments. Xylazine also induced a significant decrease in plasma insulin level within 30 min after treatments. The present data suggested that xylazine stimulates GH release in cattle.

The interactive effects of VIP, PHI, GHRH, and SRIF on the release of growth hormone from cultured adenohypophysial cells in cattle.

The effects of hypothalamic peptides [vasoactive intestinal peptide (VIP), peptide histidine isoleucine (PHI), growth hormone (GH)-releasing hormone (GHRH) and somatostatin (SRIF) on GH release from cultured bovine adenohypophysial cells were studied. The cells were incubated for 2 h with the peptides after preincubation for 3.5 days. At doses from 10(-9) to 10(-7)M VIP, the amount of GH released was significantly greater than in the controls (P < 0.05 to P < 0.001). PHI (10(-10 to 10(-7)M did not alter the bovine GH concentration in the media. Incubation with the media containing 10(-7)M GHRH, 10(-7)M VIP, and combined treatment with the VIP plus GHRH increased GH by 186, 40 and 182%, respectively (P < 0.001). Furthermore, although VIP-induced GH release was significantly decreased by SRIF compared with the treatment with VIP alone (P<0.001), the VIP significantly blunted the inhibitory effect of the SRIF on GH release by 24% when compared with that of the SRIF plus GHRH without the VIP (P < 0.05). GH release in combined treatments with VIP, GHRH and SRIF was significantly less than that of the VIP plus GHRH (P < 0.001), but it was significant 29% increase compared with the SRIF plus GHRH (P < 0.05). The combined effects of the VIP (10(-7)M) with GHRH (10(-7), 10(-8) and 10(-10)M significantly induced GH release compared with the controls (P < 0.001), but no additive effect was not observed when compared with the GHRH alone. The results indicate that VIP, but not PHI, acts directly on cultured adenohypophysial cells to induce GH release in cattle.