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Genetic profiles for 46 microsatellite loci of 12 inbred strains of rats, including 2 congenic strains and a coisogenic strain, have been demonstrated. Rates of loci with different alleles between 2 inbred strains, which are not closely related to each other in origin, were from 71.7% between ACI/N and IS/Kyo strains to 41.3% between F344/N and TM/Kyo. On the other hand, the rates were 0% in both of 2 sets of congenic strains; between F344/N and F344/N-rnu, or between BN/fMaiKyo and BN.IS. When WTC/Kyo and the coisogenic strain TRM/Kyo (WTC/Kyo-tm) were compared for 115 microsatellite loci, no loci with different alleles between the strains were found. The 46 loci should be useful as genetic monitoring markers, since all of the primer pairs generate distinct PCR-products at a fixed annealing temperature of 55 degrees C.

Spontaneously epileptic rats (SER) are a double mutant (zi/zi, tm/tm) obtained by mating the tremor rats with another tremulous mutant, zitter rats, and spontaneously exhibit absence-like seizures, tonic convulsions and wild jumping or running episodes. In addition, SER exhibit retardation in learning ability under differential reinforcement at a low rate (DRL)-5 sec schedule of food reinforcement in operant behavior at 14 to 15 weeks of age. We examined the long-term effects of phenobarbital (PB) on total bar presses, reinforcements and inter-response time distribution to assess whether the poorness of learning ability in SER can be ascribed to frequent occurrence of seizures. Food pellets containing 0.1% PB were given ad libitum to male SER from 7 weeks, and operant behavior was examined at 13 to 14 weeks of age. Tonic seizures and wild jumping or running episode were inhibited effectively and poor operant performance was significantly improved by continual intake of PB in SER.

Two genetic polymorphisms of salivary proteins were found by polyacrylamide gel electrophoresis among inbred strains of rats. Both proteins (RSP-1 and RSP-2) were inherited as a single autosomal trait. The loci were designated as Rsp-1 (rat salivary protein-1) and Rsp-2. Rsp-1 had two codominant alleles (Rsp-1a, Rsp-1b), and Rsp-2 had two alleles (Rsp-2a, and Rsp-2b); Rsp-2a was dominant over Rsp-2b. The Rsp-1 locus is not linked with the linkage groups (LGs) I, II, IV, V and the LGs containing Acp-2 and Pg-1. The Rsp-2 is not linked with the LGs I, II, V, X and the LGs containing Amy-1, Es-6 and Pg-1.

The sensitivity of spontaneously epileptic rats (SER), double homozygotes of zitter and tremor mutations, to external stimuli that induce seizures was studied in comparison with tremor (tm/tm) and zitter (zi/zi) rats, and with normal Kyo: Wistar and F 344/N rats. Touching their body, a blinking light (1200 lux, 1 sec interval) or a big sound (8 and 12 kHz, 95 dB) induced tonic extension only in the SER. The response frequency was 22 to 44% at 9 weeks of age and 75 to 100% at 13 weeks of age. Electric stimulus at 30 mA induced tonic-clonic convulsions in all Kyo: Wistar and F 344/N rats. At 20 mA the incidence of seizures decreased with age, from 100% at 5 weeks of age to 33% at 13 weeks of age in Kyo: Wistar rats and from 100 to 71% in F344/N rats. In SER, 10-mA stimuli induced tonic extension at 9 and 13 weeks of age, and 20 and 30 mA induced tonic convulsions, generalized or partial convulsions, and wild jumping or running episodes at 5, 9 and 13 weeks of age. At 30 mA, the incidence of convulsive seizures decreased with age in both tremor (tm/tm) and zitter (zi/zi) rats. Apparently external stimuli acted as simple triggers in the induction of tonic extension, since characteristic tonic extension is induced in the SER by each of the stimuli used in the present study, and induced convulsions closely resembled spontaneously occurring convulsions. The threshold of external stimuli in the induction of tonic extension became lower with aging in the SER, indicating that they are appropriate models for evaluating anticonvulsant drugs, as reported previously.

Postnatal behavioral development and learning ability of operant performance were examined in spontaneously epileptic rats (SER: zi/zi, tm/tm), and the original tremorous mutant strains of rats, tremor rats (tm/tm) and zitter rats (zi/zi) and their controls. Before the eyes opened, the increase in body weight and the age of achieving the righting reflex on a surface were no significantly different between the SER and their littermates without epileptic seizures (SER-N: zi/zi, tm/+ or zi/zi, +/+), and between tremor rats and the original strain Kyo: Wistar rats. After the eyes opened, the increase in body weight, age of achieving the righting reflex in air and traction performance, and the development of rotarod performance, were delayed in SER and tremor rats in comparison with other groups of rats. The zitter rats were apparently inferior in their development of rotarod performance in comparison with the same zitter homozygous SER-N. Operant performance was more inferior in SER than in SER-N and in tremor rats than in Kyo: Wistar rats. The differences were much more marked between SER and SER-N than between tremor and Kyo: Wistar rats. Thus, homologous tm genes and the coexistence of homologous tm and zi genes have an inhibitory effect on postnatal behavioral development and learning ability.

The low plasma alkaline phosphatase (ALP) activity of the IS strain of rats was genetically analyzed by using the cross matings with the BN strain. In the F1 hybrids, the ALP activities were intermediate but largely biased to the low activity. In the (F1 x BN) back-cross progeny, the distributions of plasma ALP activities were apparently bimodal in both sexes. Alp-2 locus with two alleles of Alp-2h and Alp-2l is tentatively proposed. Alp-2 locus did not link to the rat linkage group (LG) I, II, IV, V, and LGs containing Amy-1 and Cs-1. In the (F1 x IS) back-cross progeny, the plasma ALP activities were not different from that of the F1 hybrids and the distribution was not bimodal. Alp-2l may be dominant over Alp-2h. One third of the back-cross progeny showed vertebral anomalies and the difference of plasma ALP activities was not detected between the rats with and without vertebral anomalies.

Cramer (1981) reported that Pg-1 (urinary pepsinogen) in the rat was loosely linked with albinism in the linkage group I. We performed a three point test on the loci of pg-1, c, and Hbb. We could reconfirm that pg-1 was autosomal trait with two co-dominant alleles of pg-1a and pg-1b. But in progeny of ((WF X IS) X WF) backcross, pg-1 was linked to neither c nor Hbb, while a close linkage between c and Hbb was detected. Also pg-1 was not linked to Mup-1 (LG II), a (LG IV), Es-3 (LG V), and h (LG VI). pg-1 will be one of the most valuable genetic markers of the rats, since pg-1 was highly polymorphic among inbred strains of rats, and not linked to LG I, II, IV, V, and VI.

A micro-agglutination test method for detecting antibodies to Brucella canis was developed. Heat-killed Brucella canis antigens were diluted to an optical density of 0.8 at 420 nm using a spectrophotometer. A volume of 0.025 ml of the antigen was incubated with the same volume of serially diluted sera for 18 to 24 hr at 37 C. Titers of selected dog sera obtained by the present micro-test method were well correlated with those obtained by the classical tube test with satisfactory reproducibility. The micro-test method is more advantageous for screening the antibodies of dog sera because the test can be performed with: (1) smaller volume of the test sera and the antigen (2) shorter period for incubation, and (3) lesser labor.

Only two types of pepsinogens, which are products of the Pg-1 locus, are present in rat urine. In gastric mucosa, however, additional pepsinogen isozymes are expressed. We have found a polymorphism for rat gastric mucosa pepsinogen using agarose gel electrophoresis. Some inbred rat strains expressed a pepsinogen band, while others did not. The trait was found to be controlled by a single autosomal locus. We tentatively designated the locus as Pg-2 with two alleles, Pg-2a for the one controlling presence of the band and Pg-2o for the one controlling absence. Linkage analysis using BN and TM strains revealed that Pg-2 was closely linked to Pg-1 (3.7 +/- 1.8 cM), and that it did not belong to LG I (Hbb and p), LG II (Acon-1 and Mup-1), LG IV (Hao-1 and Svp-1), LG V (Es-1 and Es-3), LG VI (Gc and h), LG IX (RT1), LG X (Fh and Pep-3), nor a LG containing Ahd-2 (as yet undetermined).

Six male dogs orally inoculated with 3.2 X 10(8) Brucella canis and 2 male dogs naturally infected with the organisms were examined weekly for the presence of the organisms in urine and blood as well as for agglutinin titers in serum. Excretion of the organisms into urine started on 1 to 3 weeks after onset of bacteremia, i.e. 4 to 8 weeks after inoculation, and lasted for about 1 to 1.5 years. Non-bacteremic intervals, intermittent or lasting for about 1 to 6 months, followed the initial period of abundant excretion of viable organisms in urine. The highest urinary concentrations of the organisms, 2.5 X 10(4) to 1.5 X 10(6) cells per ml, were obtained in all dogs between the 6th and 14th weeks after the oral inoculation. Serum agglutinin titer rose on the 3rd to 5th weeks after the inoculation and then the titers remained at 1 : 640-1 : 2,560. The titers showed a downward trend over the 40 th to 56th weeks after the inoculation. In one of the spontaneously infected cases, the organisms were demonstrable only from urine for half a year with suspicious serum agglutinin titer of 1 : 160 after the bacteremic phase.

Electrophoresis of tear proteins on agarose gel showed polymorphism in the fastest migrating protein among 32 inbred strains of rats. In 8 strains, the protein was missing (RTP-2 B), while the other strains expressed the protein (RTP-2 A). The trait was found to be controlled by a single autosomal locus. The designation Rtp-2 locus, with two alleles (Rtp-2a, Rtp-2b), is tentatively proposed. The Rtp-2 locus is loosely linked to c locus, with a recombination frequency of 36.7 +/- 5.4 percent.

Two tester strains of the rats for testing coat color genes were established. The names, the origins and the genotypes of the rats were as follows: (1) abh (F4) Origin: selected from (BN/fMai X Kyo: Wistar) F2. Genotype: C/C, a/a, b/b, h/h. (2) bhd (F2) Origin: selected from (abh X Tester Moriyama)F2. Genotype: C/C, a/a, b/b, h/h, D/d and C/C, a/a, b/b, h/h, d/d.

In a colony of Kyo: Wistar strain of rats, we found animals with curled whiskers and hair. These rats exhibited tremor when they moved. There were no sex differences in phenotype and behavior, and the affected animals of both sexes were sterile. Among pairs that produced at least one tremulous offspring, 21.8% of the females and 21.7% of the males were affected; these proportions suggest that the anomaly is caused by an autosomal recessive gene. When the presumed heterozygous males were crossed with WAG/Rij females, about half of their female F1 hybrids were heterozygous, and they produced 26.1% (43/165) of the affected offspring when backcrossed to the sires. Again these results suggested that the disorder is caused by an autosomal recessive gene. We tentatively designated the gene as tremor (tm). The main pathological changes were seen in the gonad and central nervous system. The gonads of both sexes were aplastic even in adult animals. Vacuole formation was seen widely in the central nervous system and sometimes exhibited a spongy appearance. After administration of alpha-methyl-p-tyrosine, the norepinephrine content of the cerebellum was high, indicating that some anomalies of catecholamine release were present. The mutation is being maintained by random mating of the littermates of affected animals. Detailed pathological, endocrinological, neuropharmacological, and genetical studies are proceeding.

The allelic frequencies at 6 isozyme loci (Es-1, Es-2, Es-3, Es-4, Es-Si and Amy-1) were examined in 4 outbred Wistar strains and additionally 2 SD strains as controls. From the allelic frequencies, the averages of the heterozygosities in individual strains and the geometric genetic distances between every pair of all possible strain combinations were calculated. The averages of the heterozygosities in 2 SD strains were both intermediate (around 0.2) and the genetic distance between them was rather short. But among the Wistar strains, the averages of the heterozygosities were highly variable and the genetic relationships among them were very variable in their genetic distances. From these results, it was suggested that the outbred Wistar strains were different each other in their genetic constitutions and that no suggestion was obtained to descriminate genetically the Wistar strains from the SD strains.

In a [(BN X TM) X TM] backcross progeny of rats, nine significant linkage associations were found among 105 pairwise combinations of 15 loci. After comparing this with other published data and data of personal communications, we considered that the d gene we tentatively designated may be identical to the gene for pink-eyed dilution (p), and that the associations of Gc-Hbb, RT1-h, and Gc-Fh were due to chance rather than real linkage. The linkages obtained in this study, therefore, were Hbb-p (26.5 +/- 5.5) in LG I, Mup-1-Acon-1 (12.5 +/- 4.1) in LG II, Hao-1-Svp-1 (23.8 +/- 6.6) in LG IV, Es-1-Es-3 (17.2 +/- 4.7) in LG V, h-Gc (10.9 +/- 3.3) in LG VI, and Fh-Pep-3 (32.3 +/- 5.9) in LG X.